sign in sign up
<<
Home , Abby Dernburg, Alan Cowman, Anant Parekh, Ann Graybiel, Athene Donald, Birmingham Blitz

4 February 2005

Vol. 307 No. 5710 Pages 629–796 $10 07%.'+%#%+& 2416'+0(70%6+10 37#06+6#6+8' 51(69#4' #/2.+(+%#6+10 %'..$+1.1); %.10+0) /+%41#44#;5 #0#.;5+5 #0#.;5+5 2%4 51.76+105

Finish first with a superior . 50% faster real-time results with FullVelocity™ QPCR Kits!

Our FullVelocity™ master mixes use a novel enzyme species to deliver Superior Performance vs. Taq -Based Reagents

FullVelocity™ Taq -Based real-time results faster than conventional reagents. With a simple change Reagent Kits Reagent Kits Enzyme species High-speed Thermus to the thermal profile on your existing real-time PCR system, the archaeal Fast time to results FullVelocity technology provides you high-speed amplification without Enzyme thermostability dUTP incorporation requiring any special equipment or re-optimization. SYBR® Green tolerance Price per reaction $$$ • Fast, economical • Efficient, specific and • Probe and SYBR® results sensitive Green chemistries

Need More Information? Give Us A Call: Ask Us About These Great Products:

Stratagene USA and Stratagene Europe FullVelocity™ QPCR Master Mix* 600561 Order: (800) 424-5444 x3 Order: 00800-7000-7000 FullVelocity™ QRT-PCR Master Mix* 600562 Technical Services: (800) 894-1304 Technical Services: 00800-7400-7400 FullVelocity™ SYBR® Green QPCR Master Mix 600581 FullVelocity™ SYBR® Green QRT-PCR Master Mix 600582 Stratagene Japan K.K. *U.S. Patent Nos. 6,528,254, 6,548,250, and patents pending. Order: 03-5159-2060 Purchase of these products is accompanied by a license to use them in the Polymerase Chain Reaction (PCR) Technical Services: 03-5159-2070 process in conjunction with a thermal cycler whose use in the automated performance of the PCR process is YYYUVTCVCIGPGEQO covered by the up-front license fee, either by payment to Applied Biosystems or as purchased, i.e., an authorized thermal cycler. Use of labeling reagents may require licenses from entities other than Stratagene. SYBR is a registered trademark of Molecular Probes. Taqman is a registered trademark of Roche Molecular Systems, Inc.

GE01-05

Part of GE Healthcare

Biotrak immunoassays – the fast track to disease understanding

Cancer, heart disease, Parkinson's, multiple sclerosis, liver disease, kidney disease, inflammatory bowel disease and bowel cancer, osteoporosis, rheumatoid arthritis and wound healing. All covered.

Biotrak™ Assays from GE Healthcare are the widest available range of fully validated immunoassay kits. They give you the power to track key processes for many disease states, and assure fast, reproducible results you can rely on. To meet your needs even better, we are expanding the range; the latest Biotrak innovations include new easy-to-use ELISA kits for cancer research. These deliver significant time savings thanks to a protocol with few steps, while maintaining high sensitivity. Discover how Biotrak can power your disease research. And be sure to check back regularly for the latest developments. Visit www.amershambiosciences.com/biotrak

© 2004 Company - All rights reserved. Amersham Biosciences UK Ltd, a , going to market as GE Healthcare. OVER Meandering, rain-swollen wadis combine to form branching networks across C Volume 307 the arid landscape of southeastern Jordan. This false-color scene was acquired in May 2001 by the ASTER instrument onboard NASA’s Terra satellite. The Gordon Research 4 February 2005 Conference on Visualization in Science and Education will be held 3 to 8 July 2005 at Number 5710 Queen’s College, Oxford, UK. The schedules for the 2005 Gordon Research Conferences begin on page 746. [Image: NASA and USGS]

DEPARTMENTS 665 DEVELOPMENTAL 639 SCIENCE ONLINE The Unexpected Brains Behind Blood Vessel Growth 641 THIS WEEK IN SCIENCE 668 U.K. UNIVERSITIES 645 EDITORIAL by Patrick Bateson Desirable Scientific Conduct ‘Darwinian’ Funding and the Demise of and 646 EDITORS’CHOICE 650 CONTACT SCIENCE 670 RANDOM SAMPLES 651 NETWATCH 745 NEW PRODUCTS LETTERS 746 GORDON RESEARCH CONFERENCES 673 Versus Invention D. Premack and A. Premack. 770 SCIENCE CAREERS Elephants, Ecology, and Nonequilibrium? C. Hambler et al.;A.W. Illius. Response L. Gillson et al. National Environmental Policy Act at 35 D. A. Bronstein et al. NEWS OF THE WEEK 660 652 BIOMEDICINE BOOKS ET AL. Move Provokes Bruising Fight Over 676 EVOLUTION U.K. Biomedical Institute The Ancestor’s Tale A Pilgrimage to the Dawn of Life; 653 SPACE SCIENCE The Ancestor’s Tale A Pilgrimage to the Dawn of NASA Probe to Examine Edge of Solar System Evolution R. Dawkins, reviewed by C. F. Delwiche 653 AIDS TREATMENT A Step Toward Cheaper Anti-HIV Therapy 677 EVOLUTION Assembling the 655 QUANTUM COMPUTING J. Cracraft and M. J. Donoghue, Eds., reviewed by Safer Coin Tosses Point to Better Way for S. J. Steppan Enemies to Swap Messages 678 Browsings 655 SCIENCESCOPE

656 ESSAY Immortality Dies As Show Their Age 679 GLOBAL VOICES OF SCIENCE It Takes a Village: Medical Research 656 U.K. UNIVERSITIES and Ethics in Mali Cash-Short Schools Aim to Raise Fees, 676 O. K. Doumbo Recruit Foreign Students

657 SOUTH ASIA TSUNAMI PERSPECTIVES Powerful Tsunami’s Impact on Coral Reefs 682 OCEAN SCIENCE Was Hit and Miss The Ocean’s Seismic Hum S. Kedar and F. H. Webb 659 TAIWAN University Spending Plan 683 Triggers Heated Debate A Century of Corn Selection W. G. Hill 659 Proposed Law Targets 684 ECOLOGY -Rights Activists Untangling an Entangled Bank D. Storch, P. A. Marquet, K. J. Gaston NEWS FOCUS 686 ASTRONOMY 660 At the Heart of the Milky Way Asia Jockeys for Stem Cell Lead T. J. W. Lazio and T. N. LaRosa U.S. States Offer Asia Stiff 679 Competition 687 SIGNAL TRANSDUCTION Asian Countries Permit Research, Signaling Specificity in Yeast With Safeguards E. A. Elion, M. Qi, W. Chen

Contents continued www.sciencemag.org SCIENCE VOL 307 4 FEBRUARY 2005 633 Integrated Solutions — Analysis Real-time, multiplex PCR can be so simple —hit all targets in one tube!

Up to 4plex PCR

Detection of t(8;14) translocation sequenceinduplex PCR using the QuantiTect Multiplex PCR Kit

Detection of CSBG sequencein triplex PCR using the QuantiTect Multiplex PCR NoROX Kit

Success in quantitative, real-time, multiplex PCR at the first attempt! Get accurate results in quantitative, real-time, multiplex PCR of cDNA and genomic DNA targets without optimizing reaction and cycling conditions! Using the QuantiTect® Multiplex PCR Kit (with ROX dye) or the new QuantiTect Multiplex PCR NoROX Kit (without ROX dye), you canperform sensitive 2plex, 3plex,or 4plex PCR on appropriate real-time cyclers. Benefits of QuantiTect Multiplex PCR Kits: I Nooptimization required — reagents and protocols arepre-optimized I High sensitivity —detection of as few as 10 copies of each target sequence I Reliable quantification — target and reference genes are quantified in the same well or tube I Easy handling — ready-to-usemaster mix for use with a wide range of real-time cyclers Visit www.qiagen.com/goto/qmpcr for simple real-time multiplex PCR!

Trademarks: QIAGEN®, QuantiTect® (QIAGEN Group). PurchaseofQIAGEN products for PCR containing HotStarTaq DNA Polymeraseis accompanied by a limited license to use them in the Polymerase Chain Reaction (PCR)process for research and development activities in conjunction with a thermal cycler whose usein the automated performanceof the PCR process is covered by the up-front license fee, either by payment to Applied Biosystems or as purchased, i.e. an authorized thermal cycler. The PCR process is covered by U.S. Patents 4,683,195 and 4,683,202 and foreign equivalents owned by Hoffmann-La Roche AG. The 5' nucleaseprocess is covered by patents owned by Roche Molecular Systems, Inc. and F. Hoffmann-La Roche Ltd. Patents of thirdparties in certain countries may cover the process of multiplex PCR or of certain applications. GEXQTM1104S1WW 11/2004© 2004 QIAGEN, all rights reserved.

WWW. QIAGEN. COM www.sciencedigital.org/subscribe For just US$130, you can join A A A S TODAY and t receiving Science Digital Edition immediately! www.sciencedigital.org/subscribe For just US$130, you can join A A A S TODAY and star t receiving Science Digital Edition immediately! SCIENCE EXPRESS www.sciencexpress.org CHEMISTRY: Laser-Initiated Shuttling of a Molecule Between H-Bonding Sites J. R. Clarkson, E. Baquero, V. A. Shubert, E. M. Myshakin, K. D. Jordan, T. S. Zwier Light energy is used to move a single water molecule between two different binding sites on a single solute molecule, allowing detailed measurement of the binding energies. : RNA Polymerase IV Directs Silencing of Endogenous DNA A. J. Herr, M. B. Jensen, T. Dalmay, D. C. Baulcombe A newly described polymerase found only in plants is required for small to silence transgenes and a retroelement in Arabidopsis. CELL BIOLOGY: Chaperone Activity of Protein O-Fucosyltransferase 1 Promotes Notch Receptor Folding T. Okajima, A. Xu, L. Lei, K. D. Irvine An enzyme thought to add glucose groups to a key receptor protein as it travels to the membrane unexpectedly also acts as a chaperone to ensure correct folding of the receptor. : Insect Sex-Pheromone Signals Mediated by Specific Combinations of Olfactory Receptors T. Nakagawa, T. Sakurai, T. Nishioka, K. Touhara Receptors for insect pheromones rely on coexpression of an olfactory receptor for proper membrane insertion and for pheromone-triggered current flow.

TECHNICAL COMMENT ABSTRACTS 675 Comment on “Long-Lived with Overexpressed dFOXO in Adult Fat Body” M. Tatar full text at www.sciencemag.org/cgi/content/full/307/5710/675a Response to Comment on “Long-Lived Drosophila with Overexpressed dFOXO in Adult Fat Body” M. E. Giannakou, M. Goss, M. A. Jünger, E. Hafen, S. J. Leevers, L. Partridge full text at www.sciencemag.org/cgi/content/full/307/5710/675b BREVIA 689 MICROBIOLOGY: Simple Foraminifera Flourish at the Ocean’s Deepest Point Y. Todo, H. Kitazato, J. Hashimoto, A. J. Gooday Newly described species of tubular and round protists that thrive at depths of 10 kilometers in Pacific trenches lack calcified walls and resemble early evolutionary forms. 696 RESEARCH ARTICLES 690 STRUCTURAL BIOLOGY: Crystal Structure of a Complex Between the Catalytic and Regulatory (RIα) Subunits of PKA C. Kim, N.-H. Xuong, S. S. Taylor The structure of protein kinase C shows that cyclic AMP activates the enzyme by substituting for two amino acids of the catalytic subunit and displacing the inhibitory subunit. 696 PLANETARY SCIENCE: Saturn’s Temperature Field from High-Resolution Middle-Infrared Imaging G. S. Orton and P. A. Yanamandra-Fisher High atmospheric temperatures near Saturn’s south pole, imaged from the Keck I Telescope, probably reflect the 15-year summer in the southern hemisphere.

REPORTS 698 ATMOSPHERIC SCIENCE: Rapid Formation of Sulfuric Acid Particles at Near-Atmospheric Conditions T. Berndt, O. Böge, F. Stratmann, J. Heintzenberg, M. Kulmala Experiments show that sulfuric acid and water can react without ammonia to form new particles at a rate high enough to explain their natural atmospheric abundance. 701 : Dislocations in Complex Materials M. F. Chisholm, S. Kumar, P. Hazzledine High-resolution transmission electron microscopy confirms that many common materials deform in a complex manner by propagation of partial dislocations along two or more planes. 701 703 CHEMISTRY: End States in One-Dimensional Atom Chains J. N. Crain and D. T. Pierce Atoms at the ends of a single-atom-wide gold chain on a silicon surface have distinctive electronic states that favorably lower energy levels within the chains. Contents continued

www.sciencemag.org SCIENCE VOL 307 4 FEBRUARY 2005 635

REPORTS CONTINUED

706 GEOCHEMISTRY: Photic Zone Euxinia During the - Superanoxic Event K. Grice et al. Organic compounds and sulfur isotopes found at the Permian-Triassic boundary in Australia and China imply that was depleted in the upper ocean at that time.

709 PALEONTOLOGY: Abrupt and Gradual Extinction Among Late Permian Land Vertebrates in the Karoo Basin, P. D. Ward, J. Botha, R. Buick, M. O. De Kock, D. H. Erwin, G. H. Garrison, J. L. Kirschvink, R. Smith Correlation of sections in the Karoo Basin imply a period of enhanced vertebrate extinction before the end-Permian catastrophe, and some replacement by Triassic species.

714 : Aconitase Couples Metabolic Regulation to Mitochondrial DNA Maintenance X. J. Chen, X. Wang, B. A. Kaufman, R. A. Butow One of the proteins that packages mitochondrial DNA is a well-known metabolic enzyme, linking energy and mitochondrial DNA stability.

718 EVOLUTION: Natural Selection and Developmental Constraints in the Evolution of Allometries W. A. Frankino, B. J. Zwaan, D. L. Stern, P. M. Brakefield Artificial selection readily changes the ratio of body size to wing size in butterflies, indicating that the relative 718 sizes of body parts are shaped by selection, not developmental constraints.

720 DEVELOPMENTAL BIOLOGY: Mechanisms of Hair Graying: Incomplete Melanocyte Stem Cell Maintenance in the Niche E. K. Nishimura, S. R. Granter, D. E. Fisher Hair turns gray when stem cells in the hair follicle can no longer replenish the supply of pigment-producing cells.

724 CELL CYCLE: Dynamic Complex Formation During the Yeast Cell Cycle U. de Lichtenberg, L. J. Jensen, S. Brunak, P. Bork Only two-thirds of the proteins involved in are transcribed in a periodic fashion, but these form cell cycle protein complexes and confer periodic function to the ensemble.

727 MICROBIOLOGY: Escape of Intracellular Shigella from Autophagy M. Ogawa, T. Yoshimori, T. Suzuki, H. Sagara, N. Mizushima, C. Sasakawa Harmful bacteria disguise their identity by coating telltale surface proteins with other proteins, thereby escaping digestion by the cells they invade.

IMMUNOLOGY 731 Nod2-Dependent Regulation of Innate and Adaptive Immunity in the Intestinal Tract K. S. Kobayashi, M. Chamaillard, Y. Ogura, O. Henegariu, N. Inohara, G. Nuñez, R. A. Flavell 734 Nod2 in Crohn’s Disease Potentiates NF-κB Activity and IL-1β Processing S. Maeda, L.-C. Hsu, H. Liu, L. A. Bankston, M. Iimura, M. F. Kagnoff, L. Eckmann, M. Karin Mice lacking a gene associated with human Crohn’s disease succumb to intestinal because they 731 & have lower concentrations of antimicrobial peptides in their gut. 734 739 VIROLOGY: The Kaposin B Protein of KSHV Activates the p38/MK2 Pathway and Stabilizes Cytokine mRNAs C. McCormick and D. Ganem A protein from the herpesvirus that causes Kaposi’s sarcoma exacerbates the disease by inhibiting degradation of cytokine mRNAs in the host, increasing inflammation.

741 ECOLOGY: Mutualistic Fungi Control Crop Diversity in Fungus-Growing Ants M. Poulsen and J. J. Boomsma Fungal strains farmed by ant colonies maintain exclusivity by producing compounds that, when eaten by the ants and deposited in their manure, exclude competing fungal strains.

SCIENCE (ISSN 0036-8075) is published weekly on Friday, except the last week in December, by the American Association for the Advancement of Science, 1200 New Avenue, NW,Washington, DC 20005. Periodicals Mail postage (publication No. 484460) paid at Washington, DC, and additional mailing offices. Copyright © 2005 by the American Association for the Advancement of Science. The title SCIENCE is a registered trademark of the AAAS. Domestic individual membership and subscription (51 issues): $135 ($74 allocated to subscription). Domestic institutional subscription (51 issues): $550; Foreign postage extra: Mexico, Caribbean (surface mail) $55; other countries (air assist delivery) $85. First class, airmail, student, and emeritus rates on request. Canadian rates with GST available upon request, GST #1254 88122. Publications Mail Agreement Number 1069624. Printed in the U.S.A. Change of address: allow 4 weeks, giving old and new addresses and 8-digit account number. Postmaster: Send change of address to Science, P.O. Box 1811, Danbury, CT 06813–1811. Single copy sales: $10.00 per issue prepaid includes surface postage; bulk rates on request. Authorization to photocopy material for internal or personal use under circumstances not falling within the fair use provisions of the Copyright Act is granted by AAAS to libraries and other users registered with the Copyright Clearance Center (CCC) Transactional Reporting Service, provided that $15.00 per article is paid directly to CCC, 222 Rosewood Drive, Danvers, MA 01923. The identification code for Science is 0036-8075/83 $15.00. Science is indexed in the Reader’s Guide to Periodical Literature and in several specialized indexes. Contents continued

www.sciencemag.org SCIENCE VOL 307 4 FEBRUARY 2005 637 Western Blotting Systems and Reagents

The complete blotting solution is easy to spot.

The industry leader in innovative and powerful protein blotting equipment, Bio-Rad offers an extensive line of blotting kits, membranes, and reagents to meet all your blotting needs.

■ An array of versatile electrophoretic transfer systems for efficient transfer of proteins from a broad range of gel sizes ■ Microfiltration or dot-blotting devices for easy, reproducible binding of proteins and nucleic acids to membranes ■ HRP- and AP-based chemiluminescent and colorimetric detection kits and substrates — for all western blotting applications ■ High-quality blotting-grade reagents to simplify your blotting experience ■ A wide range of precut membranes and membrane sandwiches in nitrocellulose, supported nitrocellulose, and PVDF formats

For more information on protein blotting equipment and reagents, contact your local Bio-Rad representative or visit us on the Web at discover.bio-rad.com

Visit us on the Web at discover.bio-rad.com Call toll free at 1-800-4BIORAD (1-800-424-6723); outside the US, contact your local sales office. sciencenow www.sciencenow.org DAILY NEWS COVERAGE The Dream Difference www.scienceonline.org Socially aggressive dreams are more likely to occur during REM than non-REM sleep. New Trigger for Breast Cancer Research implicates virus-associated immune system component in the disease. Extinguished Earth What would the world look like had there been no forest fires?

science’s next wave www.nextwave.org CAREER RESOURCES FOR YOUNG SCIENTISTS

POSTDOC NETWORK: NIH Multiple-PI Policy May Open Opportunities for Postdocs B. Benderly Whether postdocs benefit from new federal policy will depend on lab and university politics. MISCINET: Creating Engineering Programs in Tribal Colleges E. Francisco With help from NSF, tribal colleges will create engineering programs leading to the bachelor’s degree. CANADA: Weeding Out the Bugs A. Fazekas Vice president of R&D at a bio-agricultural firm talks about her journey from bench to boardroom. FRANCE: French Postdocs Abroad—Finding Your Way Back Home E. Pain Next Wave speaks to two French postdocs who have left France for the and Japan. EUROPE: European Science Bytes Next Wave Staff Read the latest funding, training, and job market news from Europe. Engineering at tribal colleges. GRANTSNET: February 2005 Funding News Edited by S. Otto This is the latest index of research funding, scholarships, fellowships, and internships.

science’s sage ke www.sageke.org SCIENCE OF AGING KNOWLEDGE ENVIRONMENT

REVIEW: Lipofuscin and Aging—A Matter of Toxic Waste D. A. Gray and J. Woulfe Accumulation of this pigment might cause catastrophic lysosomal and proteasomal inhibition. NEWS FOCUS: Ageless No More M. Leslie Once thought immortal, gut bacteria suffer aging’s toll. NEWS FOCUS: Plugged Up R. J. Davenport Broken pump abets overload after a stroke.

Garbage catastrophe?

science’s stke www.stke.org SIGNAL TRANSDUCTION KNOWLEDGE ENVIRONMENT

PERSPECTIVE: Alpha Subunit Position and GABA Receptor Function D. R. Burt Experiments with linked subunits reveal the importance of subunit position to GABAA receptor function. PERSPECTIVE: Reaching Out Beyond the Synapse—Glial Intercellular Waves Coordinate Metabolism A. Charles Synaptic activity stimulates glial calcium waves, which elicit secondary sodium and metabolic waves that may produce lactate as an energy substrate for neurons. Propagation of astrocyte waves. TEACHING RESOURCE: Introduction—Overview of Pathways and Networks and GPCR Signaling R. Iyengar These lecture materials introduce general principles and emerging theory in cell signaling research.

Separate individual or institutional subscriptions to these products may be required for full-text access.

GrantsNet AIDScience Members Only! Functional www.grantsnet.org www.aidscience.com www.AAASMember.org www.sciencegenomics.org RESEARCH FUNDING DATABASE HIV PREVENTION & VACCINE RESEARCH AAAS ONLINE COMMUNITY NEWS,RESEARCH,RESOURCES

www.sciencemag.org SCIENCE VOL 307 4 FEBRUARY 2005 639 Roche Applied Science LightCycler Real-Time PCR System Insist on More Accurate Quantification of Gene Expression

Quantify more accurately with the LightCycler Instrument Efficiency Efficiency Without correction correction with ■ efficiency with linear non-linear Cycle faster to minimize non-specific products that may overestimate correction fit function fit function copy numbers. ■ Analyze all samples in the same thermal chamber to ensure temperature homogeneity and consistent PCR efficiencies.

Calibrator-normalized target/housekeeping ratios Analyze data more accurately with LightCycler Relative 40 ng 1.03 1.18 1.41 Quantification Software 8 ng 2.21 1.79 1.01 1.6 ng 6.00 4.17 1.17 ■ Use calibrator normalization to ensure consistency between PCR runs. 3.08 2.38 1.21 Mean ■ S.D. 2.60 1.58 0.22 Within runs, rely on an efficiency-correction feature that accounts for C.V. 84.3% 66.4% 18.0% differences in PCR efficiencies between target and reference genes. ■ Figure 1: Impact of different PCR efficiency adjustments Obtain sample concentrations from non-linear standard curves to on accuracy of relative quantification. Total RNA was more precisely quantify low-copy genes, which often suffer from non- used for quantitative RT-PCR on the LightCycler System. linear PCR efficiencies (Figure 1). Sample data were evaluated with the LightCycler Relative Quantification Software, using the efficiency correction Shouldn’t accurate quantification be the primary goal of gene functions described above, to generate calibrator-normalized expression studies? Contact your Roche Applied Science representative target/housekeeping ratios. The significantly lower Coefficient www.lightcycler-online.com of Variation (C.V.) demonstrates the greater accuracy made and visit today! possible by the LightCycler Software’s use of efficiency corrections and a non-linear fit function.

LightCycler is a trademark of a member of the Roche Group. Roche Diagnostics GmbH The technology used for the LightCycler System is licensed Roche Applied Science from Idaho Technology, Inc., Salt Lake City, UT, USA. 68298 Mannheim © 2005 Roche Diagnostics GmbH. All rights reserved. THIS WEEK IN edited by Stella Hurtley and Phil Szuromi

Saturnian Hot Spot Glimpses into the P/T Boundary Ground-based infrared observations of Saturn with the Long The Permian-Triassic extinction was the most extreme in Earth’s Wavelength Spectrometer on the Keck I Telescope on Mauna Kea history. It has been difficult in part to determine the environmental reveal a hot spot in the atmosphere within 3° of the south pole, a conditions that may have led to the extinction. Grice et al. (p. 706, warm polar cap, anomalous temperature bands, and oscillations in published online 20 January 2005) present a detailed chemical temperatures in the southern hemisphere that are not correlated analysis of marine sections with cloud patterns. Orton and Yanamandra-Fisher (p. 696) sug- obtained by drilling off west- gest these features are related to radiative forcing and dynamical ern Australia and South China. forcing that are consistent The data suggest that the with 15 years of constant upper part of the oceans at solar illumination of the Fungus Monoculture the time of the extinction southern hemisphere as Sat- were extremely oxygen poor urn goes through its southern on the Ant Farm and sulfide rich. Ward et al. summer solstice. Leaf-cutting ants live in obligate (p. 709, published online 20 ectosymbiosis with clonal fungi January 2005), in contrast, re- Two’s Company, that they rear for food. These construct a record of the ter- symbionts are vertically trans- restrial vertebrate extinctions Three’s a Cloud? ferred during colony foundation, in the Karoo Basin, Africa. This It has long been thought that but fungus gardens are, in princi- area preserves the most de- the in situ creation of new ple, open for horizontal sym- tailed vertebrate record (secondary) cloud condensa- biont transmission later on. from this time, but correlating tion nuclei arises mainly from Poulsen and Boomsma (p. 741) rocks in different parts of the the reaction of gas phase sul- show that fungal ectosymbionts prevent competing fungal strains Basin has been problematic. furic acid and water, but the from becoming established by ancient incompatibility mechanisms Using paleomagnetism and rate of particle formation ob- that have not been lost despite millions of years of domestication isotopes, they show served in laboratory studies and single-strain rearing by ants. These fungal incompatibility com- that extinctions were acceler- has been too slow (by many pounds travel through the ant gut to make the ant feces incompati- ated up to a pulse at the orders of magnitude) to ac- ble with unrelated strains of symbiont. Thus, the fungi manipulate boundary, and that the pattern count for the number concen- the symbiosis to their own advantage at the expense of the ants’ of appearance of Triassic fauna trations found in . A potential interest in a genetically more diverse agriculture. may imply that some originat- faster, ternary mechanism that ed even before the final pulse. includes ammonia has been postulated on the basis of theoretical factors. Berndt et al. (p. 698) now report experimental production of particles from a mixture of Protein Kinase Inhibition Revealed sulfuric acid and water at concentrations like those naturally found An important target of the second-messenger cyclic adenosine in the atmosphere, with ammonia at concentrations lower than monophosphate (cAMP) is protein kinase A (PKA). PKA, which those normally observed. The measured rate is consistent with that regulates processes as diverse as growth, memory, and metabo- required to explain atmosphere number concentrations. lism, exists as an inactive complex of two catalytic subunits and a regulatory subunit dimer. cAMP binds to the regulatory sub- units and facilitates dissociation and activation of the catalytic The End of the Line subunits. Kim et al. (p. 690) have determined the 2.0 angstrom The breaking of the translation symmetry of crystals at their sur- resolution structure of the PKA catalytic subunit bound to a dele- faces gives rise to localized surface electronic states, and, in princi- tion mutant of the regulatory subunit (RIα). The complex pro- ple, similar effects should be seen at the ends of one-dimensional vides a molecular mechanism for inhibition of PKA and suggests wires. Crain and Pierce (p. 703) present experimental evidence for how cAMP binding leads to activation. such electronic states at the ends of one-dimensional gold chains of gold grown on the stepped Si(553) A Matter of Scale surface. Scanning A striking feature of morphological diversity across animal species tunneling micro- is the variability in the relative size, or allometry, of different ap- scopy images show pendages. Virtually nothing is known of the forces that underlie markedly different the evolution of scaling relationships. Using the butterfly species contrast for the end Bicyclus anynana, Frankino et al. (p. 718) tested the roles of devel- atoms of chains opmental constraints and natural selection in determining the size when the bias volt- of the wings relative to the body, which as a measure of wing load- age is reversed, and differential conductance measurements ing has clear functional and ecological importance. Artificial selec- reveal the details of the electronic states of the end atoms that tion experiments on the size of the forewing relative to overall agree well with the results of tight-binding calculations. The for- body size resulted in a rapid evolutionary response. In this case, de- mation of end states helps lower the energy of filled states for velopmental constraints did not limit the evolution of the scaling CONTINUED ON PAGE 643

CREDITS: (TOP TO BOTTOM) POULSEN AND BOOMSMA; POULSEN CREDITS:AND PIERCE BOTTOM) TO CRAIN (TOP atoms within the chain.

www.sciencemag.org SCIENCE VOL 307 4 FEBRUARY 2005 641 What if moving from one particular protein to the most relevant journal and patent literature were as easy as pushing a button?

It is.

Not only does SciFinder provide access to more proteins and nucleic acids than any publicly available source, but they’re a single click away from their referencing patents and original research.

Coverage includes everything from the U.S. National Library of ’s (NLM) MEDLINE® and much more. In fact, SciFinder is the only single source of patents and journals worldwide. Once you’ve found relevant literature, you can use SciFinder’s powerful refinement tools to focus on a specific research area, for example: biological studies such as target organisms or diseases; expression microarrays; or analytical studies such as immunoassays, fluorescence, or PCR analysis. From each reference, you can link to the electronic full text of the original paper or patent, plus use citation tools to track how the research has evolved and been applied. Visualization tools help you understand results at a glance. You can categorize topics and substances, identify relationships between areas of study, and see areas that haven’t been explored at all. Comprehensive, intuitive, seamless—SciFinder directs you. It’s part of the process. To find out more, call us at 1-800-753-4227 () or 1-614-447-3700 (worldwide) or visit www.cas.org/SCIFINDER.

Part of the process.SM

A division of the American Chemical Society. SciFinder is a registered trademark of the American Chemical Society. “Part of the process” is a service mark of the American Chemical Society. CONTINUED FROM 641 Molecular THIS WEEK IN Genetics of relationship. Instead, it is the pattern of natural selection imposed by the external envi- Bacteria & Phages ronment that determine the wing-body size allometry. MADISON, WI • AUGUST 2-7, 2005 www.wisc.edu/meetings Packaging and Power Combining Sessions • RNA Polymerase Structure/Function Mitochondrial DNA (mtDNA) is packaged with proteins into a nucleoid. Chen et al. • Transcriptional Regulation (p. 714) show that one of the mtDNA packaging proteins is the Krebs cycle enzyme, • DNA Replication, Recombination and aconitase, that the mitochondrion uses to generate metabolic energy. In this second Transposition role, aconitase is required for mtDNA maintenance under particular metabolic condi- • Translation and Postranscriptional tions. This finding provides a direct link between energy generation and mtDNA stabili- Regulation ty, mitochondrial disease, and aging. • Global Regulation and Stress Response • Molecular Biology of • Bacterial Cell Biology Fade to Gray • Bacteriophage Development and Host Interactions Aging brings on many changes in the human • Genomics and Proteomics body, among them the graying of hair. Nishimura • Surfaces and Signaling et al. (p. 720; published online 23 December 2004) found in a mouse model of hair graying Session Chairs that a deficiency of the gene Bcl-2 caused pro- Tania Baker • MIT gressive loss of pigment cells in the bulge of the Steve Busby • Univ. of hair follicle—the hair stem-cell niche. Thus, the Carlos Catalano • Univ. of Colorado HSC physiology of hair graying involves defective self- Kenn Gerdes • University of S. maintenance of melanocyte stem cells with ag- Anna Karls • University of Georgia ing, and may serve as a paradigm for understanding aging mechanisms in other tissues. Janine Maddock • Univ. of Michigan Karen Ottemann • UC Santa Cruz Mathias Springer • CNRS Paris Autophagic Arms Race David Thanassi • SUNY Stony Brook Dmitry Vassylyev • UAB One defense against intracellular invaders is to enclose them within autophagic vac- uoles that then fuse with degradative lysosomes to destroy the . Ogawa et al. (p. 727, published online 2 December 2004) show that the invading bacterial pathogen Shigella can be recognized and trapped by autophagy. Generally, the pathogen circumvents the autophagic event by secreting an effector protein called IcsB during multiplication within the host cytoplasm; mutant bacteria lacking IcsB are par- ticularly susceptible to autophagic killing. The Shigella VirG protein acts as the target q: How can I organize that stimulates autophagy, but the IscB protein can camouflage it. and protect my back issues of Science? Giving Mice the Nod a:Custom-made The detection of bacteria in the gut by the immune system is regulated, in part, by the Nod library file cases! proteins, which recognize peptidoglycan motifs from bacteria, and there is a strong associa- Designed to hold tion of the inflammatory bowel disorder Crohn’s disease with in the Nod2 gene. 12 issues and covered in a rich Nevertheless, questions remain about the normal physiological role of the Nod proteins in burgundy leather- maintaining in the gut and how impaired Nod function leads to inflammation. like material, each et al slipcase includes Maeda . (p. 734) observed that Nod mutations in mice, corresponding with those car- an attractive label ried by Crohn’s disease patients, increased susceptibility to intestinal inflammation caused with the Science by the bacterial cell wall precursor muramyl dipeptide. Kobayashi et al. (p. 731) generated logo. Nod2-deficient mice. Although these did not spontaneously develop intestinal in- One ...... $15 Great gift idea! flammation, they were more susceptible to oral infection with the bacterial pathogen Listeria Three ...... $40 monocytogenes. Production of a group of mucosal antimicrobial peptides was particularly Six ...... $80 diminished in Nod2-deficient animals, which suggests that a similar defect may contribute Send order to: to inflammatory bowel disease in humans. TNC Enterprises Dept.SC P.O. Box 2475 Warminster, PA 18974 Specify number of slipcases and Cytokine Production and Kaposi’s enclose name, address and payment with your order (no P.O. boxes please). When tissues are infected with Kaposi’s sarcoma–associated herpesvirus (KSHV), they Add $3.50 per slipcase for shipping produce large amounts of proinflammatory cytokines that are linked to disease pro- and handling. PA residents add 6% sales tax. Cannot ship outside U.S. gression. McCormick and Ganem (p. 739) show that a viral protein, kaposin B inter- Credit Card Orders: AmEx, VISA, ET AL. acts with mitogen-activated protein kinase–associated protein kinase 2 and enhances MC accepted. Send name, number, the activity of this host cell protein, serving to block the decay of AU-rich messenger exp. date and signature. RNAs and increase the level of secreted cytokines. This result explains the association Order online: of KSHV-related disease and enhanced cytokine production. www.tncenterprises.net/sc

CREDIT: NISHIMURA Unconditionally Guaranteed

www.sciencemag.org SCIENCE VOL 307 4 FEBRUARY 2005 643 Access OptiMEF™ Primary Mouse Embryonic Fibroblasts (MEFs) From ArtisOptimus

Capture the Power of Primary MEFs. Advance your Easy Access. Recognized as the gold standard, widespread understanding of gene function. OptiMEF™ knockout and adoption of primary MEFs has been hampered by the lack of a wild-type cells are convenient tools for analysis of gene convenient source of cells. Now, using ArtisOptimus OptiMEF™ function and regulation. cells, you can avoid the cost, delays and inconvenience of Known Genetic Background. When using cell culture requesting primary MEFs from other researchers or the hassle model systems you want results you can rely on. Unlike of generating your own MEFs. immortal and transformed cell lines, OptiMEF™ cells Convenient Custom Services. Submit your mouse knockout retain their initial growth characteris- model for primary MEF production on a custom basis. tics and genetic backgrounds. Avoid Get the ArtisOptimus Advantage. Don’t monkey around, artifacts and inconclusive results access knockout, transgenic, inbred and hybrid OptiMEF™ observed with immortalized cells, cells today. use OptiMEF™ cells for biologically Accelerate relevant results. your research. www.artisoptimus.com OptiMEF™ Mdr1a/b -/- at Passage 5 1-760-918-8900 Don’t monkey around with inferior immortalized cells. EDITORIAL Desirable Scientific Conduct

any scientists are aware of the subtle influences on their own scientific conduct, but many others are not. Sydney Brenner, the joint winner of the 2002 for physiology or medicine, delightfully described a slide in which data points were scattered very close to a straight line— but a large mysterious black object lay in one corner. By degrees, the onlooker realizes that the object is a thumb placed over a data point that is far away from the straight line. The thumb covered it up because the result was supposedly anomalous. The test tube was dirty, the animal was sick thatM day, or something else was amiss. Many other such rationalizations—perhaps they should be called rules of thumb—are produced and the damage done may not serious. A famous example of selective use of the thumb was provided by the data on which Gregor Mendel based his laws of inheritance. The eminent statistician R. A. Fisher argued that, on grounds of probability, the data were too good to be true. Mendel had presumably started to see (correctly as it turned out) a pattern while he was still doing the critical breeding experiments and then began to drop data that did not fit. Another form of data selection can lead to serious error. Before the discovery of stratospheric ozone holes in the 1980s, statistical analysis of satellite data threw out the “outliers” on the assumption that such measurements were unreliable. It was only when scientists working at one station in repeatedly obtained low values that the processing mistake was discovered and the ozone holes recognized. Treasure your exceptions! The data point lying under the researcher’s thumb might be the most interesting result of the whole study. Social psychologists and sociologists have long been aware of the subtle ways in which bias can creep into research. The behavior of their subjects sometimes results not from the effects of any experimental manipulation, but merely from the attention paid to them by the experimenter. Much evidence suggests that experimenters often obtain the results they expect to obtain, partly because they unwittingly influence the outcome of the experiment.* The expectancy effect is sometimes comparable in size to the effect of the experimental manipulation itself. Many scientists take appropriate steps to avoid this kind of bias. They use “blind” procedures so that the person making the measurements cannot uncon- sciously bias the result. Analysis is carried out ideally while the researcher remains unaware of the identity of each group. Although many are careful, others are not and do not even recognize the problem. Suspect findings damage unnecessarily the reputation of scientists for integrity, lending weight to the more bizarre views about the social construction of science. The reality of prejudice or theoretical conformism in scientific work emphasizes that a considerable job of educating many members of the scientific community is still needed. That kind of awareness becomes all the more necessary when issues of funding and promotion are at stake. Some notorious cases have demonstrated just how ferocious can be the pressure from commercial funders to ignore good scientific practice. A well-known example was the shameful treatment at the University of Toronto of Nancy Olivieri, who published data uncongenial to the drug company that had funded her.† Sources of funding can undoubtedly exert corrupting influences on scientific behavior. The bad cases should be condemned when they are discovered. “Affiliation bias” may, however, be much more subtle, leading research workers to select evidence suiting their own preconceptions. All scientists need to be very careful about how evidence was obtained in the first place. Desirable modes of scientific conduct require considerable self-awareness as well as a reaffirmation of the old virtues of honesty, scepticism, and integrity. Patrick Bateson

Patrick Bateson is in the Sub-Department of Animal Behaviour, , High Street, Madingley, Cambridge CB3 8AA, UK. E-mail [email protected] *R. Rosenthal, D. B. Rubin, Behav. Brain Sci. 1, 377 (1978). †D. G. Nathan, D.Weatherall, N. Engl. J. Med. 347, 1368 (2002).

10.1126/science.1107915 SMITH Y E. TERR : CREDIT

www.sciencemag.org SCIENCE VOL 307 4 FEBRUARY 2005 645 EDITORS’ CHOICE H IGHLIGHTS OF THE R ECENT L ITERATURE edited by Gilbert Chin

rivers appear to be eroding their bedrock at the geologically Don’t Keep Hedgehogs extreme and unsustainable rates of several centimeters Rescuing a hedgehog victim of a road acci- per year; hence, other processes dent and nurturing it back to health can be must be contributing to deeply satisfying. But Riley and Chomel bedrock dynamics. Over a show that the payback from such 7-year period, Stock et al. an exotic pet may not be entirely monitored several rivers in benign. Hedgehogs harbor a Taiwan and in the Pacific variety of pathogens that are Northwest of the United potentially transferable to States.These rapidly eroding humans and our livestock. European (top) and African pygmy rivers had all been historically Several species of hedgehogs (left) hedgehogs. scoured of sediment.This history have been widely introduced and the authors’ measurements into the United States and are ago, domestication brought humans imply that long-term stream kept illegally in some states, in into contact with a range of new erosion, at least in areas with the extraordinary number of 40,000 pathogens; the current vogue for exotic weak bedrock, is influenced households.A recent survey shows that they pets and food animals will do likewise, more by the ability of rivers can carry foot-and-mouth disease virus, namely, monkeypox and in prairie to entrain a thin covering of Salmonella, and Mycobacteria, as well as dogs and SARS in civets. —CA sediment, which reduces wear, dermal fungal .Thousands of years Emerg. Infect. Dis. 11, 1 (2005). than specific bedrock properties. In areas of high slope, debris flows, which periodically CELL BIOLOGY they present evidence that rophore until it is cleaved by scour streams and rivers and Popeye’s Ribosomes this explains why Fe-S cluster esterases within a cell.They thus allow rapid downcutting, biogenesis is an essential coupled rhodamine 110 to may be the most critical Ribosomes are the central function and thus why two o-hydroxycinnamic acid factor. — BH component of the protein mitochondria (where Fe-S derivatives via amide linkages. Geol. Soc.Am. Bull. 117, 174 (2005). synthesis machinery, and cluster biogenesis originates) In their acetylated form,

the efficient manufacture are essential. — SMH these side chains force the BEHAVIOR . (2005). 10.1021/JA043736V of ribosomes is crucial. These EMBO J. 10.1038/sj.emboj.7600540; rhodamine core to adopt a Less Editing, Less machines contain roughly 10.1038/sj.emboj.7600541 (2005). nonfluorescing lactonized 60 protein and RNA parts. configuration. Ester cleavage Depression The assembly of these parts CHEMISTRY causes the side chains to form A number of recent studies occurs in the nucleus and Unlocking a hydrocoumarin, which is have fueled a sense of ET AL.,J.AM. CHEM. SOC involves importing proteins Fluorescence favored by steric interaction of optimism that the fuzzy link from the cytosol and placing the methyl groups, and liberates between genes and behavior them onto ribosomal RNA The power of fluorescent the fluorescent acid form of might be firmed up and (rRNA); subsequently, the probes can be enhanced by rhodamine 110.The authors made explicit, an especially assembled small and large controlling how and when the followed the uptake of the challenging task given the ribosomal subunits are probe becomes excitable. latent fluorophore into HeLa likelihood that the contribu- exported to the cytosol. Chandran et al. describe an cells, where they observed tions of individual genes Two groups describe an approach for masking fluores- strong fluorescence from the (and distinct mutations) to unanticipated link in this cence from a xanthene fluo- cytosol and lysosomes but not behavior might be only a few chain of events—iron-sulfur from the nucleus. — PDS percent of the total mix of (Fe-S) cluster biosynthesis. J.Am. Chem. Soc. 10.1021/ja043736v predisposition, motivation, H H (2005). In a screen for ribosomal -O N O N O- and environment. Biogenic O O export mutants,Yarunin et al. O amines are, of course, front found that genes implicated O GEOLOGY and center in any consideration in Fe-S cluster biosynthesis Protection Against of mood and affect, and genes in the cytosol are needed for rhodamine 110 + 2 Erosion encoding various aspects of ribosomal export. In particular, serotonin function in neurons + the protein Rli1 requires a H2N O NH2 River and stream erosion rates (synthesis, transport, and

Fe-S cluster to promote rRNA AcO OH and the resulting river profiles receptors) have already been -O2C processing and small ribosomal O are becoming more widely rec- targeted as prime candidates subunit export. Kispal et al. ognized as depending on a for dysfunction in depression. also implicate Rli1 in the The intramolecular rearrange- complicated and incompletely Englander et al. have used export of ribosomal subunits ments that liberate rhodamine documented suite of factors. a pair of mice strains to

from the nucleus. In addition, (left). For instance, some streams and examine the interaction of CREDITS:ARNOLD, BUCHELE/WWI / PETER RUPERT (TOP) INC., LEFT) JOE MCDONALD/CORBIS; (TOP CHANDRAN (BOTTOM)

646 4 FEBRUARY 2005 VOL 307 SCIENCE www.sciencemag.org        serotonin receptors, stressful situations, detailed global estimates of its distribution    and a selective serotonin reuptake and magnitude. Measuring NPP from inhibitor (SSRI).They report that, in space has failed to provide convincing    comparison to C57BL/6 mice, BALB/c values, because two essential parameters, animals have lower serotonin levels phytoplankton carbon biomass and a  (due to a polymorphism in tryptophan term related to the physiological status      hydroxylase-2) and are generally easier of the organisms, are not directly to stress (via a behavioral despair task). quantifiable remotely.     Furthermore, the type 2C serotonin Behrenfeld et al. start with satellite          receptor in BALB/c mice undergoes less measurements of the chlorophyll content of        editing of its pre-messenger RNA, and upper ocean and the backscattering this yields, in compensatory fashion, of certain wavelengths of light (which they receptors that are more sensitive to use to estimate phytoplankton carbon serotonin.Administering the despair biomass), and then estimate phytoplankton task or the SSRI (the antidepressant growth rates and fluoxetine) bumps up the extent of RNA calculate NPP. editing and presumably titrates down- They can do ward the responsiveness of postsynaptic this by taking neurons to released serotonin.The advantage of unexpected finding is that this change in laboratory editing due to drug or stress is not seen studies that if both are given together, suggesting have shown that the molecular response may be that the ratio influenced by the state of the subject of chlorophyll and blocked by antidepressants. — GJC to carbon bio- J. Neurosci. 25, 648 (2005). mass is a calcula- ble function of EARTH SCIENCE Getting a Fix on Fixation Phytoplankton growth rates during the boreal summer (top) and winter (bottom).      Marine net primary production (NPP)         is a measure of how much atmospheric changes in light, nutrients, and temperature.          carbon is fixed via by This work brings nearer the prospect of      organisms in the ocean. Until now, producing a more accurate picture of global         only direct field sampling has yielded marine NPP over space and time. — HJS          accurate estimates of NPP, which has           Global Biogeochem. Cycles 19,         severely limited attempts to obtain 10.1029/2004GB002299 (2005).                  

H IGHLIGHTED IN S CIENCE’ S S IGNAL T RANSDUCTION K NOWLEDGE E NVIRONMENT           Balancing Axons and Dendrites       Neurons are polarized cells with axons (signal output) and    dendrites (signal input). Not only are these functionally    distinct parts of the cell, but they differ in morphology too.    Jiang et al. report that when glycogen synthase kinase 3β (GSK-3β) activity    was increased by transfection of isolated embryonic hippocampal neurons with a constitutively active mutant, the number of cells that formed an axon decreased, β      and when GSK-3 activity was inhibited, the number of cells producing multiple     axons increased even though the overall number of neurites did not change.    They identified the phosphatidylinositol 3-kinase (PI3K) pathway as a stimulator  

, 10.1029/2004GB002299 (2005). β β    19 of GSK-3 phosphorylation, which results in an inhibition of GSK-3 . Activation     of the PI3K pathway by expression of the kinase Akt or inactivation of the   phosphatase PTEN produced multiaxon neurons. Yoshimura et al. show that   GSK-3β phosphorylates collapsin response mediator protein 2 (CRMP-2), which is known to contribute to axon formation.Treatment of neurons with neurotrophin      3 (NT-3) or brain-derived neurotrophic factor (BDNF) stimulated axon growth      and decreased CRMP-2 phosphorylation. Furthermore, the stimulation in axon      length was blocked if CRMP-2 abundance was decreased.Thus, a pathway involving

ET AL., GLOBAL BIOGEOCHEM. CYCLES    PI3K regulates the activity of GSK-3β and the phosphorylation of the assembly regulatory protein CRMP-2, and hence controls axon formation         and growth in neurons. —NG                           Cell 120, 123; 137 (2005).                     CREDITS: BEHRENFELD                                  www.sciencemag.org SCIENCE VOL 307 4 FEBRUARY 2005      Who’s taking science to new heights?

Science is essential reading on the way to the top. It takes several days to reach the top in big wall climbing, so you can only carry the bare essentials. When you calculate the information content to weight ratio, is there any more concentrated reading source than Science? “ AAAS member Dr. R. Douglas Fields, senior scientist, developmental neuroscience ” ouglas Fields

.D

.R

Dr

Photo:

Douglas Fields’ son Dylan takes a break on the way up

AAAS is committed to advancing science and giving a To join the international family of science, go to voice to scientists around the world. We work to improve www.aaas.org/join. science education, promote a sound science policy, and support human rights. Helping our members stay abreast of their field is a key priority for AAAS. One way we do this is through Science, which features all the latest breakthroughs and ground- breaking research, and keeps scientists connected wherever they happen to be. Members like Douglas find it essential reading. www.aaas.org/join www.sciencemag.org EDITOR-IN-CHIEF Donald Kennedy EXECUTIVE PUBLISHER Alan I. Leshner EXECUTIVE EDITOR Monica M. Bradford PUBLISHER Beth Rosner DEPUTY EDITORS NEWS EDITOR R. Brooks Hanson, Katrina L. Kelner Colin Norman FULFILLMENT & MEMBERSHIP SERVICES ([email protected]) DIRECTOR Marlene Zendell; FULFILLMENT SYSTEMS: MANAGER Waylon EDITORIAL SUPERVISORY SENIOR EDITORS Barbara Jasny, Phillip D. Szuromi; Butler; MEMBER SERVICES: MANAGER Michael ; SENIOR SPECIALIST Pat SENIOR EDITOR/PERSPECTIVES Orla Smith; SENIOR EDITORS Gilbert J. Chin, Butler; SPECIALISTS Laurie Baker, Tamara Alfson, Karena Smith, 1200 New York Avenue, NW Pamela J. Hines, Paula A. Kiberstis (), Beverly A. Purnell, L. Bryan Andrew Vargo; MARKETING ASSOCIATE Deborah Stromberg Ray, Guy Riddihough (Manila), David Voss; ASSOCIATE EDITORS Lisa D. BUSINESS OPERATIONS AND ADMINISTRATION DIRECTOR Deborah Rivera- Washington, DC 20005 Chong, Marc S. Lavine, H. Jesse Smith, Valda Vinson, Jake S. Yeston; Wienhold; BUSINESS MANAGER Randy Yi; SENIOR FINANCIAL ANALYSTS Lisa Editorial: 202-326-6550, FAX 202-289-7562 ONLINE EDITOR Stewart Wills; ASSOCIATE ONLINE EDITOR Tara S. Marathe; BOOK Donovan, Jason Hendricks; ANALYST Jessica Tierney, Farida Yeasmin; News: 202-326-6500, FAX 202-371-9227 REVIEW EDITOR Sherman J. Suter; ASSOCIATE LETTERS EDITOR Etta Kavanagh; RIGHTS AND PERMISSIONS: ADMINISTRATOR Emilie David; ASSOCIATE Elizabeth INFORMATION SPECIALIST Janet Kegg; EDITORIAL MANAGER Cara Tate; SENIOR Sandler; MARKETING: DIRECTOR John Meyers; MEMBERSHIP MARKETING MANAGER Bateman House, 82-88 Hills Road COPY EDITORS Jeffrey E. Cook, Harry Jach, Barbara P. Ordway; COPY EDITORS Darryl Walter; MARKETING ASSOCIATES Karen Nedbal, Julianne Wielga; Cynthia Howe, Sabrah M. n’haRaven, Jennifer Sills, Trista Wagoner, Cambridge, UK CB2 1LQ RECRUITMENT MARKETING MANAGER Allison Pritchard; ASSOCIATES Mary Ellen Alexis Wynne; EDITORIAL COORDINATORS Carolyn Kyle, Beverly Shields; +44 (0) 1223 326500, FAX +44 (0) 1223 326501 Crowley, Amanda Donathen, Catherine Featherston; DIRECTOR OF PUBLICATION ASSISTANTS Chris Filiatreau, Joi S. Granger, Jeffrey Hearn, Lisa INTERNATIONAL MARKETING AND RECRUITMENT ADVERTISING Deborah Harris; Johnson, Scott Miller, Jerry Richardson, Brian White, Anita Wynn; INTERNATIONAL MARKETING MANAGER Wendy Sturley; MARKETING/MEMBER SUBSCRIPTION SERVICES For change of address, missing issues, EDITORIAL ASSISTANTS Ramatoulaye Diop, E. Annie Hall, Patricia M. SERVICES EXECUTIVE: Linda Rusk; JAPAN SALES AND MARKETING MANAGER Jason new orders and renewals, and payment questions: 800-731- Moore, Brendan Nardozzi, Jamie M. Wilson; EXECUTIVE ASSISTANT Hannaford; SITE LICENSE SALES:DIRECTOR Tom Ryan; SALES AND CUSTOMER SERVICE 4939 or 202-326-6417, FAX 202-842-1065. Mailing addresses: Sylvia S. Kihara; ADMINISTRATIVE SUPPORT Patricia F. Fisher AAAS, P.O. Box 1811, Danbury, CT 06813 or AAAS Member Mehan Dossani, Catherine Holland, Adam Banner,Yaniv Snir; ELECTRONIC NEWS SENIOR CORRESPONDENT Jean Marx; DEPUTY NEWS EDITORS Robert Services, 1200 New York Avenue, NW, Washington, DC 20005 MEDIA: INTERNET PRODUCTION MANAGER Lizabeth Harman; ASSISTANT PRODUCTION Coontz, Jeffrey Mervis, Leslie Roberts, John Travis; CONTRIBUTING EDITORS MANAGER Wendy Stengel; SENIOR PRODUCTION ASSOCIATES Sheila Mackall, NSTITUTIONAL ITE ICENCES Elizabeth Culotta, Polly Shulman; NEWS WRITERS Yudhijit Bhattacharjee, I S L please call 202-326-6755 for any Lisa Stanford; PRODUCTION ASSOCIATE Nichele Johnston; LEAD APPLICATIONS Jennifer Couzin, David Grimm, Constance Holden, Jocelyn Kaiser, questions or information DEVELOPER Carl Saffell REPRINTS Ordering/Billing/Status 800-635-7171; Corrections Richard A. Kerr, Eli Kintisch, Andrew Lawler (New ), Greg 202-326-6501 Miller, Elizabeth Pennisi, Charles Seife, Robert F. Service (Pacific NW), PRODUCT ADVERTISING ([email protected]); MIDWEST Rick Erik Stokstad; Amitabh Avasthi (intern); CONTRIBUTING CORRESPONDENTS Bongiovanni: 330-405-7080, FAX 330-405-7081 • WEST COAST/W. CAN- PERMISSIONS 202-326-7074, FAX 202-682-0816 Marcia Barinaga (Berkeley, CA), Barry A. Cipra,Adrian Cho, Jon Cohen ADA B. Neil Boylan (Associate Director): 650-964-2266, FAX 650- MEMBER BENEFITS Bookstore:AAAS/BarnesandNoble.com bookstore (San Diego, CA), Daniel Ferber, Ann Gibbons, Robert Irion, Mitch 964-2267 • EAST COAST/E. CANADA Christopher Breslin: 443-512-0330, www.aaas.org/bn; Car purchase discount: Subaru VIP Program Leslie (NetWatch), Charles C. Mann, Evelyn Strauss, Gary Taubes, FAX 443-512-0331 • UK/SCANDINAVIA/FRANCE/ITALY/BELGIUM/NETHERLANDS 202-326-6417; Credit Card: MBNA 800-847-7378; Car Rentals: Ingrid Wickelgren; COPY EDITORS Linda B. Felaco, Rachel Curran, Sean Andrew Davies (Associate Director): +44 (0)1782 750111, FAX +44 Hertz 800-654-2200 CDP#343457, Dollar 800-800-4000 Richardson; ADMINISTRATIVE SUPPORT Scherraine Mack, Fannie Groom (0) 1782 751999 • GERMANY/SWITZERLAND/ Tracey Peers #AA1115; AAAS Travels: Betchart Expeditions 800-252-4910; BUREAUS: Berkeley, CA: 510-652-0302, FAX 510-652-1867, New (Associate Director): +44 (0) 1782 752530, FAX +44 (0) 1782 Life Insurance: Seabury & Smith 800-424-9883; Other Benefits: England: 207-549-7755, San Diego, CA: 760-942-3252, FAX 752531 JAPAN Mashy Yoshikawa: +81 (0) 33235 5961, FAX +81 (0) AAAS Member Services 202-326-6417 or www.aaasmember.org. 760-942-4979, Pacific Northwest: 503-963-1940 33235 5852 Jessica Nachlas +9723 5449123 • TRAFFIC MANAGER [email protected] (for general editorial queries) PRODUCTION DIRECTOR James Landry; SENIOR MANAGER Wendy K. Shank; Carol Maddox; SALES COORDINATOR Deiandra Simms ASSISTANT MANAGER Rebecca Doshi; SENIOR SPECIALISTs Vicki J. Jorgensen, [email protected] (for queries about letters) CLASSIFIED ADVERTISING ([email protected]); U.S.: SALES Jessica K. Moshell, Amanda K. Skelton; SPECIALIST Jay R. Covert DIRECTOR Gabrielle Boguslawski: 718-491-1607, FAX 202-289- [email protected] (for returning manuscript reviews) REFLIGHT DIRECTOR MANAGER P David M. Tompkins; Marcus Spiegler 6742; INTERNET SALES MANAGER Beth Dwyer: 202-326-6534; INSIDE [email protected] (for book review queries) ART DIRECTOR Joshua Moglia; ASSOCIATE ART DIRECTOR Kelly Buckheit; SALES MANAGER Daryl Anderson: 202-326-6543; WEST COAST/MIDWEST Published by the American Association for the Advancement of ILLUSTRATOR Katharine Sutliff; SENIOR ART ASSOCIATES Holly Bishop, Kristine von Zedlitz: 415-956-2531; EAST COAST Jill Downing: 631- Science (AAAS), Science serves its readers as a forum for the Laura Creveling, Preston Huey, Julie White; ASSOCIATE Nayomi 580-2445; LINE AD SALES Emnet Tesfaye: 202-326-6740; SENIOR SALES presentation and discussion of important issues related to the Kevitiyagala; PHOTO RESEARCHER Leslie Blizard COORDINATOR Erika Bryant; SALES COORDINATORS Rohan Edmonson, advancement of science, including the presentation of minority or SCIENCE INTERNATIONAL Caroline Gallina, Christopher Normile, Joyce Scott, Shirley Young; conflicting points of view, rather than by publishing only material EUROPE ([email protected]) EDITORIAL: INTERNATIONAL MANAGING INTERNATIONAL: SALES MANAGER Tracy Holmes: +44 (0) 1223 326525, on which a consensus has been reached. Accordingly, all articles FAX +44 (0) 1223 326532; SALES Christina Harrison, Gareth Stapp; published in Science—including editorials, news and comment, EDITOR Andrew M. Sugden; SENIOR EDITOR/PERSPECTIVES Julia Fahrenkamp- SALES ASSISTANT Helen Moroney; JAPAN: Jason Hannaford: +81 (0) 52 and book reviews—are signed and reflect the individual views of Uppenbrink; SENIOR EDITORS Caroline Ash, Stella M. Hurtley, Ian S. the authors and not official points of view adopted by the AAAS Osborne, Peter Stern; ASSOCIATE EDITOR Stephen J. Simpson; EDITORIAL 777 9777, FAX +81 (0) 52 777 9781; PRODUCTION: MANAGER Jennifer or the institutions with which the authors are affiliated. SUPPORT Emma Westgate; ADMINISTRATIVE SUPPORT Janet Clements, Phil Rankin; ASSISTANT MANAGER Deborah Tompkins; ASSOCIATE Amy Marlow, Jill White; NEWS: INTERNATIONAL NEWS EDITOR Eliot Marshall DEPUTY Hardcastle; SENIOR TRAFFICKING ASSOCIATE Christine Hall; SENIOR PUBLI- AAAS was founded in 1848 and incorporated in 1874. Its mission is CATIONS ASSISTANT Robert Buck; PUBLICATIONS ASSISTANT Natasha Pinol to advance science and innovation throughout the world for the NEWS EDITOR Daniel Clery; CORRESPONDENT Gretchen Vogel (Berlin: +49 (0) benefit of all people. The goals of the association are to: foster 30 2809 3902, FAX +49 (0) 30 2809 8365); CONTRIBUTING AAAS BOARD OF DIRECTORS RETIRING PRESIDENT, CHAIR ; communication among scientists, engineers and the public; CORRESPONDENTS Michael Balter (Paris), Martin Enserink (Amsterdam PRESIDENT ; PRESIDENT-ELECT Gilbert S. Omenn; enhance international cooperation in science and its applications; and Paris); INTERN Mason Inman TREASURER David E. Shaw; CHIEF EXECUTIVE OFFICER Alan I. Leshner; BOARD promote the responsible conduct and use of science and technology; ASIA Japan Office: Asca Corporation, Eiko Ishioka, Fusako Tamura, 1-8- Rosina M. Bierbaum; John E. Burris; John E. Dowling; Karen A. foster education in science and technology for everyone; enhance 13, Hirano-cho, Chuo-ku, Osaka-shi, Osaka, 541-0046 Japan; +81 (0) Holbrook; Richard A. Meserve; Norine E. Noonan; Peter J. Stang; the science and technology workforce and infrastructure; increase 6 6202 6272, FAX +81 (0) 6 6202 6271; [email protected] JAPAN NEWS Kathryn D. Sullivan; Lydia Villa-Komaroff public understanding and appreciation of science and technology; BUREAU: Dennis Normile (contributing correspondent, +81 (0) 3 3391 and strengthen support for the science and technology enterprise. 0630, FAX 81 (0) 3 5936 3531; [email protected]); CHINA REPRESENTATIVE Hao Xin, + 86 (0) 10 6307 4439 or 6307 3676, FAX +86 (0) 10 6307 INFORMATION FOR CONTRIBUTORS 4358; [email protected]; SOUTH ASIA Pallava Bagla (contributing See pages 135 and 136 of the 7 January 2005 issue or access correspondent +91 (0) 11 2271 2896; [email protected]); CENTRAL ASIA www.sciencemag.org/feature/contribinfo/home.shtml Richard Stone (+7 3272 6413 35, [email protected])

ENIOR DITORIAL OARD S E B William Cumberland, UCLA Michael J. Lenardo, NIAID, NIH Thomas Stocker, Univ. of Bern John I. Brauman, Chair, Stanford Univ. , Norman L. Letvin, Beth Israel Deaconess Medical Center Jerome Strauss, Univ. of Pennsylvania Med. Center Richard Losick, Harvard Univ. Judy DeLoache, Univ. of Virginia Richard Losick, Harvard Univ. Tomoyuki Takahashi, Univ. of Tokyo Robert May, Univ. of Oxford Robert Desimone, NIMH, NIH Andrew P. MacKenzie, Univ. of St. Andrews Glenn Telling, Univ. of Kentucky Marcia McNutt, Monterey Bay Aquarium Research Inst. John Diffley, Cancer Research UK Raul Madariaga, École Normale Supérieure, Paris Marc Tessier-Lavigne, Genentech Linda Partridge, Univ. College Dennis Discher, Univ. of Pennsylvania Rick Maizels, Univ. of Edinburgh Craig B. Thompson, Univ. of Pennsylvania Vera C. Rubin, Carnegie Institution of Washington , Cancer Research UK , Brandeis Univ. Michiel van der Klis, Astronomical Inst. of Amsterdam Christopher R. Somerville, Carnegie Institution , Univ. of Geneva George M. Martin, Univ. of Washington Derek van der Kooy, Univ. of Toronto , WHO Virginia Miller, Washington Univ. , Johns Hopkins Richard Ellis, Cal Tech Edvard Moser, Norwegian Univ.of Science and Technology BOARD OF REVIEWING EDITORS , Fritz-Haber-Institut, Berlin Elizabeth G. Nabel, NHLBI, NIH Christopher A. Walsh, Douglas H. Erwin, Smithsonian Institution Naoto Nagaosa, Univ. of Tokyo Christopher T. Walsh, Harvard Medical School R. McNeill Alexander, Univ. Barry Everitt, Univ. of Cambridge James Nelson, Stanford Univ. School of Med. Graham Warren, Yale Univ. School of Med. Richard Amasino, Univ. of Wisconsin, Madison Paul G. Falkowski, Rutgers Univ. Roeland Nolte, Univ. of Nijmegen Fiona Watt, Imperial Cancer Research Fund Kristi S. Anseth, Univ. of Colorado , Univ. of Copenhagen Eric N. Olson, Univ. of Texas, SW Julia R. Weertman, Northwestern Univ. Cornelia I. Bargmann, Univ. of California, SF Barbara Finlayson-Pitts, Univ. of California, Irvine Erin O’Shea, Univ. of California, SF Daniel M. Wegner, Brenda Bass, Univ. of Utah Jeffrey S. Flier, Harvard Medical School Malcolm Parker, Imperial College Ellen D. Williams, Univ. of Maryland Ray H. Baughman, Univ. of Texas, Dallas Chris D. Frith, Univ. College London , Imperial College R. Sanders Williams, Stephen J. Benkovic, Pennsylvania St. Univ. R. Gadagkar, Indian Inst. of Science Josef Perner, Univ. of Salzburg Ian A. Wilson, The Scripps Res. Inst. Michael J. Bevan, Univ. of Washington Mary E. Galvin, Univ. of Delaware Philippe Poulin, CNRS Jerry Workman, Stowers Inst. for Medical Research Ton Bisseling, Wageningen Univ. Don Ganem, Univ. of California, SF David J. Read, Univ. of John R. Yates III,The Scripps Res. Inst. Peer Bork, EMBL John Gearhart, Johns Hopkins Univ. Colin Renfrew, Univ. of Cambridge Martin Zatz, NIMH, NIH Dennis Bray, Univ. of Cambridge Jennifer M. Graves, Australian National Univ. JoAnne Richards, Baylor College of Medicine Walter Zieglgänsberger, Max Planck Inst., Munich Stephen Buratowski, Harvard Medical School Christian Haass, Ludwig Maximilians Univ. , Univ. of Cambridge , Baylor College of Medicine Jillian M. Buriak, Univ. of Alberta Dennis L. Hartmann, Univ. of Washington Edward M. Rubin, Lawrence Berkeley National Labs Joseph A. Burns, Cornell Univ. Chris Hawkesworth, Univ. of Bristol David G. Russell, Cornell Univ. Maria Zuber, MIT William P. Butz, Population Reference Bureau Martin Heimann, Max Planck Inst., , Mass. General Hospital , Univ. of Dundee James A. Hendler, Univ. of Maryland , Institut Pasteur Mildred Cho, Stanford Univ. Ary A. Hoffmann, La Trobe Univ. Dan Schrag, Harvard Univ. BOOK REVIEW BOARD David Clapham, Children’s Hospital, Boston Evelyn L. Hu, Univ. of California, SB Georg Schulz, Albert-Ludwigs-Universität David Clary, Oxford University Meyer B. Jackson, Univ. of Wisconsin Med. School Paul Schulze-Lefert, Max Planck Inst., Cologne David Bloom, Harvard Univ. J. M. Claverie, CNRS, Marseille Stephen Jackson, Univ. of Cambridge Terrence J. Sejnowski, The Salk Institute Londa Schiebinger, Stanford Univ. Jonathan D. Cohen, Princeton Univ. Bernhard Keimer, Max Planck Inst., Stuttgart George Somero, Stanford Univ. Richard Shweder, Univ. of Robert Colwell, Univ. of Connecticut Alan B. Krueger, Princeton Univ. Christopher R. Somerville, Carnegie Institution , MIT , Royal Botanic Gardens, Kew Antonio Lanzavecchia, Inst. of Res. in Biomedicine Joan Steitz, Yale Univ. Ed Wasserman, DuPont F. Fleming Crim, Univ. of Wisconsin Anthony J. Leggett, Univ. of , Urbana-Champaign Edward I. Stiefel, Princeton Univ. Lewis Wolpert, Univ. College, London

650 4 FEBRUARY 2005 VOL 307 SCIENCE www.sciencemag.org NETWATCH edited by Mitch Leslie

LINKS Math From the Ground Up Researchers who specialize in the foundations of mathematics delve into deep topics such as set theory and proof theory to ensure that math rests on solid philosophical ground.For a comprehensive take on DATABASE the subject, visit this site from Alexander Sakharov, a computer scien- tist and Web developer in Boston. Sakharov has arranged more than Planet Earth Checkup 50 links into chapters to create a virtual textbook, with writings from Whether you’re interested in the number of threatened plant mathematicians, computer scientists, and other experts. The contri- species in various countries, changes in glacier mass, airborne butions—which include books, articles, and entries in references such lead levels, or the use of ozone-depleting compounds such as as the Stanford Encyclopedia of Philosophy—probe topics like mathe- methyl bromide, check out the Global Data Portal from the U.N. matical logic and axioms. Environment Programme. The site lets you download data on sakharov.net/foundation.html more than 450 economic and ecological variables or render RESOURCES them as a map, graph, or table. The chart above, for example, Taking the Sting Out of depicts the change in forest cover for different countries Bumblebee between 1990 and 2000, with They may be as close to cuddly as insects can Brazil showing the biggest loss get, but bumblebees give taxonomists and China recording gains. The headaches because different species inhabit- figures collected here provide ing the same area often look alike. For help the underpinnings for the U.N.’s navigating the group’s treacherous taxon- Global Environment Outlook, omy, make a beeline for this site from ento- an occassional report on the mologist Paul Williams of the Natural His- biosphere’s condition, and other tory Museum in London. His checklist of summaries. world bumblebees—the first published since geodata.grid.unep.ch 1923—attempts to tidy up the nomenclat- ural mess. You can find out which types of IMAGES bees live in North America and Europe or search the site by bioregions, such as eastern Mineral Asia. The checklists discuss valid and invalid names for each kind of bee.To aid identification, the entries also include photos of the male bees’ Mother Lode genitalia, a key feature for differentiating species.Above, a male Bombus asiaticus prowls for a mate. These glittering crystals of www.nhm.ac.uk/entomology/bombus/index.html roselite (below) owe their crim- son hue to cobalt, which consti- tutes about 10% of their weight. Find out much more about DATABASE roselite—from its chemical composition to the origin of its name—at Webmineral, an exhaustive database main- All Together Now tained by Houston, Texas–based geol- To deduce a protein’s function, researchers need to know every- ogy consultant David Barthelmy. thing from its structure and location in the cell to what molecules Since NetWatch’s last visit (Science, it interacts with.But this information resides in disparate databases 11 June 1999, p. 1731), this com- that often use different terminology, and compiling it “can be a pendium of 4300 minerals has painful experience,” says computer scientist Golan Yona of Cornell added photos for more than half University. So Yona and his colleagues crafted Biozon, a database the entries and Java applets that let that merges the holdings of more than a dozen molecular biology you study each crystal’s structure collections, including SwissProt, KEGG, PDB, and BodyMap.The site from multiple angles.You’ll also find data lets you run searches that span different data types, such as finding such as the minerals’ hardness rating, x-ray dif- 3D structures for all proteins that interact with the protein BRCA1, fraction values, classification according to the Strunz and Dana sys- which is implicated in some breast cancers.The ability to compare tems, and other tidbits. For example, roselite isn’t named for its ruddy results from different databases side by side also makes it easier to color, but for Gustav Rose, a 19th century German mineralogist. spot discrepancies. webmineral.com biozon.org

Send site suggestions to [email protected]: www.sciencemag.org/netwatch CREDITS (TOP TO BOTTOM): UNEP; JOHN VEEVAERT; PAUL WILLIAMS/NATURAL HISTORY MUSEUM BOTTOM): HISTORY TO WILLIAMS/NATURAL UNEP; CREDITS (TOP VEEVAERT; PAUL JOHN

www.sciencemag.org SCIENCE VOL 307 4 FEBRUARY 2005 651 This Week PAGE 656 657 Aging Corals bacteria after the NEWS tsunami

BIOMEDICINE among different research groups “would best be achieved through colocation with a university” with “the widest possible range Move Provokes Bruising Fight of disciplines.” The two NIMR scientists who sat on the Over U.K. Biomedical Institute task force signed off on this report—Steven Gamblin, head of the protein structure group, CAMBRIDGE, U.K.—For more than a year, cedes that moving NIMR from Mill Hill and Robin Lovell-Badge, head of develop- researchers at a world-class biomedical insti- may “cost more than it will save.” mental genetics. But both now disagree with tution in Britain have been battling to stop Blakemore cites several reasons for want- MRC’s interpretation. They say they support what they see as a clumsy and destructive ing to move NIMR despite the cost. He told the the broad conclusions but not Blakemore’s attempt to overhaul their community. Next science committee that government recom- views on relocating to London. As they tell it, week, Parliament will give its view of their mendations dating back to 1996 and earlier the consensus wanted a move to the city if this appeal to block a relocation from the suburbs have urged that the institute be brought closer proved better than staying at Mill Hill. But in to the center of London, possibly followed by to a university. And in a telephone interview, he their view, MRC has not really compared the a clear decision from their top governing spoke of MRC’s long-held concern that NIMR options in detail and is ignoring the condi- board, the Medical Research Council (MRC). is too isolated from academic and clinical life. tional clause. Lovell-Badge also told the par- Both sides say this fight may It must change to survive, he liamentary committee that in a phone call leave bruises that will affect and others argue. Blakemore had pressured him to drop his biomedical research in the A July 2004 MRC task resistance, but that he refused. Blakemore United Kingdom for years. force on the renovation plan, acknowledged that there was a conversation Leading scientists at the which included two NIMR sci- but denied that he intended any threat. National Institute for Medical entific leaders, concluded that Gamblin, Lovell-Badge, O’Garra, and oth- Research (NIMR)—the largest the institute’s “long-term suc- ers argue that NIMR already enjoys many of U.K. biomedical unit, with a cess” will depend on its ability the good things that the move to London might direct government budget of bring, such as interdiscipli- $62 million—took an angry nary collaborations and protest to the halls of Parlia- partnerships with clinics. ment in December. In the par- They offer piles of docu- liamentary inquiry that began ments as proof. But they that same month, they blasted concede the obvious: that an MRC plan to move their entire facility NIMR is not physically from its perch on a green ridge northwest of near a teaching hospital or London to a university site in the city. The university. On the other staff’s main concern, according to NIMR hand, they say, it’s doubtful immunologist Anne O’Garra and others who any London university can spoke with Science, is that the advantages of provide a secure, spacious the present spot in Mill Hill will be lost— animal facility like theirs. including a secure animal facility and an Gamblin and Lovell-Badge unparalleled 19-hectare campus—with no voiced a widely held suspi- commensurate gain. cion that the university- MRC’s preferred scheme for “renewing” dominated MRC wants to NIMR, says Colin Blake- Auction? , the U.K. biomedical research funding direc- sell Mill Hill to have more more, MRC’s chief executive, is to sell the tor, defends a plan to sell off the 19-hectare Mill Hill facility and move flexibility in the budget for entire NIMR estate. Director John Skehel the staff into the city center. academic projects—and to has announced that he will retire in 2006, subject NIMR staffers to but the scientific staff would be kept intact. to do “translational research”: adapting basic the rigors of university life. The cash from the sale, as Blakemore biology to medical uses. NIMR supports more Blakemore says he has heard the rumors, explained in the 1 December hearing before than a dozen fields of research and is known but they are “completely unfounded.” There is the House of Commons Science and Tech- for its excellence in infections and immunity, “no hidden agenda,” he adds: “If the MRC had nology Committee, would help pay for new developmental biology, neuroscience, and wanted to close down the NIMR, they would facilities in central London. The final structural biology, among others. But it has no have done it” long ago and not have “spent two details have not been set, but an MRC task clinical center and no degree-granting func- and a half years reviewing it.” force has singled out Kings College and tion. The task force found that NIMR could Parliament’s science committee will offer University College as candidate sites. It’s best make a “cultural shift” from its focus on its view of the controversy in a report to be not clear whether the government would laboratory work to clinical practice “through issued on 8 February. And Blakemore says own the new city buildings or pay a univer- physical proximity to a teaching hospital.” MRC hopes to issue its own decision on

sity to maintain them. Yet Blakemore con- And it found that encouraging crosstalk 10 February. –ELIOT MARSHALL BOTTOM): PHOTOGRAPHY;TO NIMR BARKER EVANS CREDITS (TOP

652 4 FEBRUARY 2005 VOL 307 SCIENCE www.sciencemag.org Focus 660 665 668 Asia’s Multipurpose Abandoning stem cell guidance physics and challenge chemistry

SPACE SCIENCE NASA Probe to Examine Edge of Solar System Despite its current budget troubles, NASA and expensive trip beyond Pluto. Instead, by Nuclear Spectroscopic Telescope Array, last week laid out plans to launch a mission flying in an Earth orbit beyond the magneto- designed to detect black holes—has been to explore the edge of the solar system. The sphere, the spacecraft’s two neutral-atom delayed on technical grounds. The project, by $134 million probe, slated a team at the California for a 2008 launch, will also Institute of Technology in serve President George W. Pasadena, will not get a Bush’s exploration vision green light until at least next by examining galactic cos- year, NASA officials say, mic rays that pose hazards because the design needs to humans traveling beyond further study. And tight Earth’s orbit. funding, combined with the Southwest Research In- recent reorganization of stitute of San Antonio, Texas, NASA’s science effort, tem- will lead the mission, called porarily threw planetary the Interstellar Boundary researchers for a loop last Explorer (IBEX). Chosen week. On 24 January, Curt from more than three dozen Niebur, discipline scientist proposals, IBEX is part of for the outer planets research NASA’s Small Explorer program, announced that $5 effort designed to put rela- million for data analysis had tively low-cost probes into been “redirected” and that orbit more quickly than the “the future of the program is agency’s usual space science far from secure.” But 3 days missions. “This is an excit- later, acting director of ing and breakthrough exper- Outer limits. Earth-orbiting IBEX satellite will map the region where particles from NASA’s solar system divi- iment for NASA to spon- the sun meet the tenuous currents of interstellar space. sion Andrew Dantzler wrote sor,” says Ghassem Asrar, researchers that “the funding the agency’s deputy science chief. No other imagers can detect particles that will enable has not been ” but simply moved to mission has attempted to chart the to map the boundary between the another area “for purely administrative rea- heliopause—the bubblelike transition zone solar system and deep space. sons.” Dantzler blamed the mix-up on a where the solar wind breaks down—in such Although IBEX appears set to proceed, “miscommunication” within NASA. detail. IBEX won’t actually make the long another Small Explorer mission—the –ANDREW LAWLER

AIDS TREATMENT A Step Toward Cheaper Anti-HIV Therapy The U.S. Food and Drug Administration Aspen Pharmacare of South Africa began heads the Campaign for Access to Essential (FDA) approved three generic anti-HIV the application process in September (Science, run by Médecins Sans Frontières, drugs last week, a decision that finally allows 8 October 2004, p. 213). On 25 January, argues, however, that FDA approval is unnec- U.S. government–sponsored programs to 12 days after FDA received Aspen’s completed essary. The World Health Organization offer these cheaper pills to infected people application, it “tentatively” approved three of (WHO) has already “prequalified” many who live in poor countries. its drugs. (The tentative designation means generic AIDS drugs that are in an even easier- The Bush Administration insisted last year that, for patent reasons, the drugs can be sold to-use formulation—three pills mixed into one that the so-called President’s Emergency Plan for use only in poor countries.) “I’m very fixed dose—and U.S. insistence on additional for AIDS Relief—which plans to spend pleased,” says , head of the approval by FDA “creates confusion and has $15 billion over 5 years—could only use National Institute of Allergy and Infectious wasted time and money,” says ’t Hoen. “To cel- drugs approved by FDA. Howls of protests Diseases. “This is what we were hoping would ebrate this FDA approval as a major break- followed, as many AIDS advocates and clini- happen.” The approved Aspen drugs include a through is presenting a false picture.” cians worried that this would rule out use of pill that combines AZT and 3TC, and, sepa- According to a WHO report released on cheap treatments now popular in many poor rately, nevirapine. “I hope that a lot of other 26 January, anti-HIV drugs currently reach countries. The Administration promised to companies follow suit, and we can get the ball only 700,000 of the nearly 6 million poor process completed applications from generic rolling with these generics,” says Fauci. people in the world who most urgently need

CREDIT: NASA/JPL-CALTECH COURTESY manufacturers within 2 to 6 weeks. Ellen ’t Hoen, a lawyer based in Paris who treatment. –JON COHEN

www.sciencemag.org SCIENCE VOL 307 4 FEBRUARY 2005 653 20th ANNUAL New England Biolabs Molecular Biology Summer Workshops

We are pleased to announce the twentieth annual Molecular Biology Summer Workshops, sponsored by New England Biolabs in conjunction with Smith College. Workshops are held at the Clark Science Center, Smith College, Northampton, MA, USA. Over 2,500 people have graduated from this intensive training program in the past nineteen years. Learn Molecular Biology in 2 Weeks! This intensive, two-week course emphasizes hands-on molecular biology laboratory work and covers a wide variety of topics and techniques. when: Topics/Techniques: Session 1: June 12 – June 25, 2005 :: gene cloning (cDNA and genomic) Session 2: July 10 – July 23, 2005 Session 3: July 31 – August 13, 2005 :: gene expression analysis :: PCR and quantitative RT-PCR where: :: genomics and Clark Science Center Smith College :: DNA sequencing and DNA fingerprinting Northampton, MA USA :: and much more – visit our website for a complete list to apply: Application Information: apply online at http://www.science.smith.edu/neb No previous experience in molecular biology is required or expected. Fifty participants per session will be selected from a variety of disciplines or and academic backgrounds. Mail a recent resume and one paragraph explaining your interest to: FEE: $3900 per participant includes lab manual, use of all equipment and supplies, and room and board (all rooms are singles). Molecular Biology Summer Workshops Dr. Steven A. Williams APPLICATION DEADLINE: March 31, 2005. Clark Science Center Payment in full is due by April 29, 2005. Late applications will be accepted! Smith College Your application should include a recent resume and one paragraph explaining Northampton, MA 01063 your reasons for taking the course. Please specify the session to which you are applying (1, 2, or 3) and indicate a second choice from one of the other sessions.

apply For additional information, please call (413) 247-3004 online! or visit the Summer Workshop web site: http://www.science.smith.edu/neb the leader in enzyme technology QUANTUM COMPUTING ScienceScope Safer Coin Tosses Point to Better Way Florida Rejects For Enemies to Swap Messages Chiropractic Program Alice and Bob are finally splitsville. After four possible states, two of which represent Florida’s Board of Governors has killed a pro- years of sending encrypted messages to “heads” and two of which represent “tails.” posal to set up a chiropractic school at each other, they’re getting a divorce. This changes the state of the other entangled Florida State University (FSU).The board’s 10–3 vote last week against the proposal, They’ve moved away from each other and photon, which is sent to Bob. Bob measures developed by FSU after the state legislature only communicate electronically, but this his photon, but because quantum ambiguity endorsed a $9-million-a-year spending plan, creates a problem. “They want to decide makes different pairs of the four states look caps months of protests by faculty members who’s going to keep the dog, so they toss a the same, he’s unable to determine whether who viewed the school as a threat to the coin,” says Alipasha Vaziri, a at the Alice picked heads or tails. He calls the university’s scientific reputation (Science, National Institute of Standards and Technol- flip—tells Alice heads or tails—and then 14 January, p. 194). ogy in Gaithersburg, Maryland. How to do a Alice reveals which of the four states she “There’s no way that the program can fair coin flip over a telephone wire? picked, allowing Bob to verify instantly now be resurrected,” says FSU Provost Physicists have now shown how, using whether he won or lost the toss. Lawrence Abele.“I’m glad we don’t have to quantum computers. In drag the faculty through a long and pro- an upcoming issue of tracted discussion” over the scientific merits Physical Review Letters, of chiropractics, he says. Before voting, board Vaziri and his colleagues members said they expected that a new pri- describe an experiment vate college near Daytona Beach would pro- in which Alice and Bob duce more chiropractors than Florida needs. perform a fair coin flip The board’s decision comes as a “big quantum-mechanically; relief,” says Raymond Bellamy, director for if one party tries to cheat, surgery at FSU’s Tallahassee campus,who the deception is quickly helped organize faculty opposition. Besides revealed, something that hurting FSU’s standing as a research univer- sity, he says, the proposed school “would scientists don’t know have been a horrible waste of taxpayer how to guarantee with money.” –YUDHIJIT BHATTACHARJEE classical computers. The fair electronic Europe, U.S. Differ on Mercury coin flip is what cryp- At odds. Is it possible to make a fair flip with a hidden coin, untrustwor- tographers term a thy opponents, and no referee? Quantum physics says yes. The European Commission is proposing an “post–Cold War” proto- international initiative to phase out mercury col. In standard cryptography, two parties Bob can’t cheat, because he doesn’t know production and prevent surpluses from who trust each other attempt to sneak a mes- the outcome of the flip before transmitting his flooding the world market.The new strategy, sage by an untrustworthy opponent; here, two guess to Alice. And it’s a subtler point, but released this week, would permanently shutter a major mine in Spain and ban all parties who don’t trust each other try to Alice can’t cheat because the signature of the exports by the , the world’s ensure that the other isn’t cheating. So a good coin flip is inscribed in Bob’s photon; if she largest supplier of mercury, starting in 2011. coin flip protocol should ensure that Alice, tries to lie, then this deception will likely The proposal will be discussed later this who wants to keep the dog, can’t cheat Bob. show up as “noise,” nonsensical data when month at a meeting of the United Nations With classical computing, coin flips are Bob interprets his measurements of the pho- Environment Programme in Nairobi, Kenya. tricky. There’s no provably secure way for ton. Although there’s only a certain probabil- In contrast, the United States is mulling Alice to flip the coin and have Bob call the ity of catching Alice each time she cheats, it’s the idea of countries forming voluntary toss—and ensure that one party or the other probabilistically guaranteed that she’ll be partnerships to assess the problem of mer- can’t cheat when determining the winner. caught if the ownership of the dog is deter- cury. But environmentalists say that (They could sidestep the problem by having mined by 1000 coin flips. approach won’t reduce either demand or a trusted third party do the coin toss for “The big problem in the paper was to supply. –ERIK STOKSTAD them. However, the only two-party classical come up with cheating algorithms,” says algorithms rely on mathematical constructs Zeilinger, who adds that Alice would have a Italy Pulls Out of Global Fund that nobody is certain are tamper-proof.) very hard time gaming the system. “We feel Italy has decided to withhold a promised With quantum computing, though, the that our procedure is very safe.” contribution of $130 million this year to the picture changes. Vaziri, along with physicist The experiment is important “because coin Global Fund, a partnership of private and of the University of Vienna tossing is a task in bigger protocols, such as public agencies devoted to fighting AIDS, and other physicists, exploited the quantum- multiparty communications,” says Andris , and .The cut is part of a mechanical property of “entanglement” to Ambainis, a physicist at the University of $325 million reduction in government assis- ensure that neither side can cheat. In their Waterloo in Canada. “When you have suffi- tance to nonprofits this year due to a tight setup, Alice has a pair of photons (created by ciently strong coin tossing, you multiply what economic climate. Mariangela Bavicchi, a an argon- laser shot at a barium-borate you can do securely” over communications spokesperson for the fund, calls the decision crystal) that are entangled: Measure one and lines, whether the parties trust each other or an example of “regrettable behavior at the you instantly affect the other’s properties. not, he says. And given the torrid lives of Alice international level.” Alice tosses the “coin” by forcing one and Bob, cryptographers will likely stay busy –MARTA PATERLINI

CREDIT: BRAND X PICTURES/GETTY IMAGES photon’s angular momentum to take one of for many years to come. –CHARLES SEIFE

www.sciencemag.org SCIENCE VOL 307 4 FEBRUARY 2005 655 N EWS OF THE W EEK MICROBIOLOGY Immortality Dies as Bacteria Show Their Age “I don’t want to achieve immortality possible for microbes that divide into identi- the older pole of the parent grew 2.2% more through my work,” Woody Allen once said. cal-looking daughter cells. slowly than those that inherited a younger “I want to achieve it by not dying.” To test that assumption, microbiologist pole. The bigger the difference in age, the big- Although that’s unlikely for Allen and Eric Stewart of INSERM in Paris tracked the ger the difference in growth rate, they report other higher organisms, many biologists fate of individual Escherichia coli cells. The in the February PLoS Biology—a result the believed that immortality was possible for rod-shaped bacterium divides in half to form team attributes to “decreased metabolic effi- microbes. Now, however, a new study sug- two identical-looking daughter cells, which ciency.” The results mean that even “an appar- gests that bacteria get old, a find- contain one old end, or pole, inherited from the ently symmetrically dividing organism is sub- ing that may give scientists a new parent and one new pole. When those ject to aging” and “make it unlikely that natu- tool to understand aging. “It’s one daughter cells split, only two of the ral selection produced an immortal organ- of those exciting results that four resulting cells will ism,” Stewart says. makes you take a fresh look have from Leonard Guarente, who studies aging at at what you think the original cell. the Institute of Technology, you know,” says agrees, saying that the results “put the onus gerontologist Inevitable of proof on anyone who claims that cells can Thomas Kirk- decline. Young be immortal.” wood of the E. coli (blue) Biogerontologist George Martin of the University of Newcastle in grow faster and pro- University of Washington, Seattle, calls , U.K. duce more offspring the results “conceptually very important” Biologists already knew that than old E. coli (red). but cautions that the cells that slow and when it comes to aging, all cells stop reproducing may just be taking a are not created equal. In the 1970s, Current thinking assumed break to repair themselves. If further study Kirkwood offered the disposable-soma the- that all four cells were the same. shows that the older cells are actually ory: Cells of the body, or soma, can deterio- The INSERM team tracked the growth of dying off, it would “make biogerontolo- rate, but the germ line cells have to take bet- single cells and their descendants on a spe- gists take seriously the notion that there’s ter care of themselves because they give rise cially designed microscope slide, taking aging in bacteria.” And if E. coli does get to sperm and eggs. Simple organisms such images every 2 to 4 minutes for up to 6 hours. old, researchers could use it to study “how as budding yeast engage in a subtler divi- Ultimately, they tracked 94 colonies, consist- aging occurs and how it’s regulated,” sion of labor; aging yeast parents invest ing of more than 35,000 individual cells. By Guarente adds, allowing them to get “right their freshest components in their buds. But comparing 7953 pairs of sister cells, the to the molecular heart of the matter.” biologists believed that immortality was researchers discovered that cells that inherited –DAN FERBER

U.K. UNIVERSITIES Cash-Short Schools Aim to Raise Fees, Recruit Foreign Students

CAMBRIDGE,U.K.—Even Britain’s top research Ruth Collier, the university will take advantage Oxford’s alumni make annual donations, universities say they’re broke. Although they of a new law next year that will allow variable compared to 40% to 60% of rivals’ alumni. receive regular government subsidies (see tuition charges for U.K. students up to about Oxford’s student body has been expanding p. 668), the law limits what they can charge $5662 a year. The university also hopes to at 1.5% per year, while the academic staff has students for tuition. (The rate is about $2170 recruit more foreign students, who pay many “remained static,” according to the report, and per year at present.) In addition, compared to times the domestic rate. Collier says the moti- between 1979 and 1999, the ratio of students U.S. institutions, they get only modest gifts vation is not to raise funds, because that would to teachers “deteriorated from 9:5 to 13:2.” from alumni and philanthropies. The result is bring in an additional $4.7 million per year— The plan aims to reverse that trend. It also “chronic underfunding,” says a strategic plan “a tiny proportion” of the annual revenue. calls for boosting the fraction of foreign stu- released last week by the University of Rather, the goal is to make the university more dents from 7% to 12%. But it won’t be easy. Oxford.* The problem is growing worse, competitive in the world market. Oxford and other top universities are according to the document put out by Other U.K. universities are doing like- already under pressure from the government Oxford’s vice chancellor John Ford on 24 Jan- wise, including the nation’s largest, the Uni- to increase the fraction of students they accept uary, and “radical” changes are needed, versity of Manchester, which disclosed sim- from U.K. state schools. Student and political including higher student fees. ilar plans to raise tuition fees and recruit for- leaders, meanwhile, are lobbying against uni- Currently, Oxford’s income is running eigners in January. versity plans that might make admissions eas- about $38 million per year below expenditures, Although Oxford counts itself among ier for foreign students. National Union of according to Ford’s strategic planning paper, the world’s top five research universities, Students President Kat Fletcher decried an and “nearly all of the university’s core activities the paper notes, its “fundraising efforts … approach, as she told the Guardian, that treats lose money.” The pale in comparison with those of the lead- “international students as simply pound signs helps reduce the deficit by transferring at least ing U.S. universities.” Oxford raised about that will solve the funding shortfall.” $23 million per year to the university. To help $110 million overall in 2002–03, the paper The Oxford plan has gone out to students, slow the leakage, says Oxford spokesperson points out, while in that period Harvard and faculty, and others for comment until mid- Stanford raised about $495 million and March; after that, it will be considered for a * www.ox.ac.uk/gazette/2004-5/supps/strategy.pdf $472 million, respectively. Only 5% of final decision. –ELIOT MARSHALL TIMMERMANN/INSERM AND STEFANIE CREDIT: ERIC STEWART

656 4 FEBRUARY 2005 VOL 307 SCIENCE www.sciencemag.org SOUTH ASIA TSUNAMI ScienceScope Powerful Tsunami’s Impact on Coral NIH Bans Industry Consulting Reefs Was Hit and Miss Responding to an uproar last year over indus- Early surveys suggest that coral reefs around good condition,” says Plathong. Only about try consulting by staff, the National Institutes the Indian Ocean survived December’s 15% of the area’s coral was severely damaged. of Health this week announced a ban on all such interactions by NIH intramural scien- tsunami in better shape than many had feared. David Obura of CORDIO East Africa—a tists. Many staffers will also have to sell their In the sites where researchers have looked, collaborative coral research program in the stock in biotech and drug companies. “only a few areas were severely damaged, and Indian Ocean—in Mombasa, Kenya, has NIH took a hard look at its consulting the rest should recover rapidly in the next 5 to similarly good news about the African coast. policies, which were loosened in 1995, after 10 years,” says Clive Wilkinson, a marine sci- “We were generally surprised at the [small] a December 2003 report in the Los Angeles entist with the Australian Institute of Marine and very patchy damage to the coral reef Times suggested improprieties and Congress Science in Cape Ferguson. In some places, communities,” he says. investigated. Last year, NIH Director Elias divers are already helping that recovery with Although turbid water prevented local Zerhouni proposed new limits, including a restoration efforts. government and academic divers from look- 1-year ban on all consulting.This week he In the immediate aftermath of the ing at six of the 10 sites in the Seychelles followed through by releasing an interim 26 December tsunami, Wilkinson and others selected for a preliminary assessment, the sur- regulation that will implement the ban until feared the worst. The wave’s awesome power, vey indicated that only 13% of the coral further notice.The policy does not restrict as well as sediment, pollutants, and debris colonies were damaged. NIH employees from receiving some pay- washed onto the reefs when the wave The reefs that were affected suffered dif- ments for teaching, writing, or editing. retreated, posed major threats, says Russell ferent levels of damage, some repairable and Meanwhile, the inspector general of Brainard, a U.S. National Oceanic and some not. Corals were toppled over, some- the Department of Health and Human Atmospheric Administration oceanographer times covered with sand and rubble. In some Services is investigating the conduct of based in Honolulu. If eroded mud and silt places, meter-high sea fans were pummeled Trey , an Alzheimer’s disease buried a reef, they could destroy the corals. and knocked off their perches. Several reefs researcher at the National Institute of Off the coast of , however, many were littered with debris—logs, beach beds, Mental Health. Sunderland is said to have received more than $500,000 from Pfizer reefs were spared. In January, volunteers and towels, palm trees, boat engines, and beach since 1999 without first asking for academic and government marine scientists umbrellas. These wave-driven objects approval or reporting the income. “become like bulldozers,” says Sunderland has accepted a job at Albert Brainard. “They severely Einstein College of Medicine in New York City erode the coral habitat.” but has not left NIH yet. His lawyer, Robert After the tsunami, Plathong Muse of Washington, D.C., declined to com- realized he had to act fast to ment, but he has told NIH that its “indiffer- save any damaged corals. He ence” was why Sutherland failed to file the brought a brigade of 136 vol- necessary paperwork. unteer divers to some of the –JOCELYN KAISER worst places in the Similan Islands. The divers worked to Call for Global right corals and were able to Biodiversity Agency salvage those that hadn’t slid beyond reach down the sloping PARIS—Researchers from around the world sea floor. They propped up sea have endorsed a call by French President Jacques Chirac for a new international fans—a temporary fix until organization for biodiversity research—akin Uprooted. The tsunami left some reefs untouched, but in many they could return with marine to the Intergovernmental Panel on Climate places—as shown above—it knocked down corals of all shapes cement. They also removed Change (IPCC)—that would sift through the and sizes. debris, although heavy objects science and identify priorities for nations.An had to be left behind. The IPCC-like agency could provide the field with took to the water for a first look. They evalu- repair efforts benefited in one way from the a stronger, unified voice, says Michael Loreau ated 175 sites in the Andaman Sea along the tsunami’s power: The wave was so strong that of Pierre and University in Paris, west coast of Thailand, rating each according potentially lethal silt and mud washed far out who chaired the scientific committee of a to the degree of impact. Half or more of the to sea in many areas. UNESCO meeting held here last week. coral was missing from 13% of the sites, Yet there were places “where the reefs were The 1500 scientists and politicians says Thamasak Yeemin, a marine scientist just planed off and stripped to bare rock,” says attending the meeting had little to cele- from Thailand’s Ramkhamhaeng University Wilkinson. In another part of Thailand, three of brate.The loss of species continues in Bangkok. About 40%, though, seemed four reefs surveyed were decimated. In some apace, and a 2002 goal of achieving a untouched. places, divers measured 5 millimeters of sedi- “significant reduction” in the rate of bio- Also last month in Thailand, Sakanan ment on top of the corals. The reefs off the diversity loss by 2010 appears doomed. Plathong, a marine biologist at Prince of Tamil Nadu cost in Southeast India also appear Besides more science, “we also need Songkla University in Had Yai, and 60 assis- to be severely damaged, as was coral off the action—now,” says Loreau. tants armed with cameras spent 3 weeks comb- Andaman and Nicobar Islands. “It will take –MARTIN ENSERINK ing a smaller area, the Similan Islands, one of some time before we can build a proper picture the country’s best dive spots. “In general most of the ecological ramifications of this disaster,”

CREDIT: KEVIN MOSES/AP PHOTO Similan islands that are dive sites are still in says Wilkinson. –ELIZABETH PENNISI

www.sciencemag.org SCIENCE VOL 307 4 FEBRUARY 2005 657    

                      ! !      "             !!              # #$       %     & ' (    )  *   "

+            ",    %   %   (             "

        "  "!-  "     " 

./#'012 &! 3! 3"         4        ! & '5"6   )$3*$        ! & ''')" 2 & !   (  0  7& (  3"8"  && "!  

 N EWS OF THE W EEK TAIWAN University Spending Plan Triggers Heated Debate

TOKYO—Taiwan has adopted a resources to your best, you won’t be $1.6 billion plan to strengthen its able to compete on the world stage,” research universities, reigniting a says , an astronomer who debate over how much of the in 2002 left the University of Cali- money should go to a handful of fornia, Berkeley, to become presi- leading institutions. A decision dent of National Tsinghua Univer- rests with a new cabinet now being sity in Hsinchu. The new funding assembled. scheme has increased speculation On 20 January legislators voted that Tsinghua may merge with its to allocate $315 million a year for neighbor, National Chiao Tung 5 years to refurbish university facil- University, creating an institution ities and boost faculty salaries. large enough to occupy the first tier. Under rules set out by the previous Chiang Wei-ling, vice president cabinet, most of the money would of National Central University, go to schools with 25,000 students Greener outlook? National Taiwan University would likely benefit from says he and his colleagues at or more, with the goal of turning the new spending plan. smaller schools recognize the need them into world-class universities. to give greater funding to a few top To be eligible, the schools would also need to and one-eighth that of the National University universities. “But it shouldn’t be a binary, win- take steps to become private, not-for-profit of Singapore. The survey “confirmed the or-lose situation,” says Chiang. “You have to institutions—part of a broader campaign to shocking disparity among the institutions,” have a pyramid, with some concentration at the streamline the government that also includes says Chen Teh-hua, director of the ministry’s top but enough incentive so other universities reducing the 100-plus universities on the island. Department of Higher Education. don’t get discouraged.” A 2003 report from a Taiwan currently has only two institutions of The money, expected to begin flowing in committee of top academics and government that size—National Taiwan University in Taipei June, will be parceled out in a competitive leaders recommended a 60–40 split. and National Cheng Kung University in Tainan. process. But the real fight will be over whether But the proper balance isn’t the only The rest of the money would be spent on the criteria that will govern the competition issue. The chair of that committee, former research centers affiliated with a dozen or so will be revised following a December setback National Central University president C. H. universities with active research programs. to the ruling Democratic Progress Party in par- Liu, says that privatizing higher education The Ministry of Education has published liamentary elections that has forced a reshuf- “is extremely controversial and is opposed statistics showing that Taiwan is not keeping fling of President Chen Shui-bian’s cabinet. by both government and opposition legisla- pace with its neighbors in supporting its lead- On one side are those who say Taiwan’s best tors.” Liu and others are hoping that the cab- ing universities. Per capita spending at the flag- chance of moving up the global academic lad- inet shakeup will give the government a ship National Taiwan University was one-12th der is by concentrating resources. “Higher edu- chance to rethink the entire policy. the amount at the University of Tokyo, it noted, cation is a competitive sport. If you don’t give –DENNIS NORMILE

UNITED KINGDOM Proposed Law Targets Animal-Rights Activists A new law could send animal-rights activists in of campaigns and increase pro- Britain to jail for up to 5 years if they cross the tection for companies that work line between peaceful protest and harassment with organizations that do ani- or intimidation. The new rules, introduced in mal research. Last summer, work Parliament on 31 January as an amendment to stopped at a new research lab in a larger anticrime bill, would specifically out- Oxford after construction com- law campaigns that target businesses that pro- pany shareholders received vide supplies or services to research organiza- threatening letters from activists tions. It would also make it illegal to protest (Science, 23 July 2004, p. 463). “outside someone’s home in such a way that And in recent months, animal- causes harassment, alarm, or distress to resi- rights activists have been charged dents,” according to a government statement. with dozens of attacks—includ- Out of bounds? A new law would impose harsh penalties for Barbara Davies of RDS (formerly the ing desecration of a family activists who cause economic harm to researchers or companies. Research Defense Society) in London, a grave—against a guinea pig farm lobby group that defends animal research, said near Birmingham that supplies research labs. of residences could allow police to shut the law could provide important support. But some animal-rights activists are wor- down protests at university research facili- “There have been amendments to laws on ried that the measures go too far. The law ties that happen to be near residential build- harassment and intimidation before, and we could potentially target peaceful protests and ings, he worries. “Like fishermen, if you thought that would work. But it’s getting boycotts, says Andrew Tyler of Animal Aid cast a wide net, you catch many nontarget worse,” she says. Key advantages of the new in Tonbridge, Kent, which has organized species,” he says. Parliament is expected to law, she says, are that it would make it eas- demonstrations against the Oxford lab. The debate the measure this spring.

CREDITS (TOP TO BOTTOM): COURTESY NATIONAL TAIWAN UNIVERSITY PRESS;TAIWAN SHAC BOTTOM): TO NATIONAL COURTESY CREDITS (TOP ier for authorities to charge the organizers prohibition against demonstrations in front –GRETCHEN VOGEL

www.sciencemag.org SCIENCE VOL 307 4 FEBRUARY 2005 659 News Focus

Less encumbered by societal restrictions on embryonic stem cells, scientists in the developing countries of Asia are giving Western researchers a run for their money Asia Jockeys for Stem Cell Lead

Veterinarian Woo Suk Hwang and gynecolo- schemes, and providing incentives for pri- and a culture of secrecy among scientists gist Shin Yong Moon leapt from obscurity to vate companies to join the effort. Giving hamper progress. Perhaps most pressing, scientific stardom last February when they these efforts a further boost, the region also says South Korea’s Hwang, the entire region isolated embryonic stem (ES) cells from has legions of lab workers willing to log suffers from a dearth of experienced senior cloned human cells, a world first and a key long hours, and increasing numbers of scientists to run the new programs. step toward therapeutic, or research, cloning. expatriate scientists are returning home to Coming from a region that rarely pro- work in the flourishing environment. A series of firsts duces scientific headlines, the announce- With all these advantages, Asia’s scien- Asian scientists have been at the forefront of ment by the Seoul National University tists believe that they can be fully competi- research on cloning and stem cells since its (SNU) pair stunned researchers around the tive in, and perhaps even lead, the race to har- inception. At China’s Shandong University, world. But it was no fluke. Hwang has a long ness stem cells. “Asia has never dominated embryologist Tong Dizhou produced the track record of successful animal world’s first cloned vertebrate, an cloning. Moon is South Korea’s Asian carp, in 1963. He went on to leading expert in assisted repro- create the first interspecies clone ductive technology. The duo were in 1973, by inserting European able to draw on the expertise of a carp DNA into an Asian carp egg. dozen co-authors at six institu- But Tong’s work remained almost tions. And when Western scien- unknown outside China. tists got their first peek into the Two decades later, in 1994, SNU lab, they were astounded to Ariff Bongso, an in vitro fertiliza- see state-of-the-art facilities— tion (IVF) expert at the National and an enviable supply of University of Singapore, reported egg donors. the first isolation of human ES Largely below the radar cells in the journal Human Repro- screen, the emerging economies duction. But Bongso was unable of South Korea, Singapore, Tai- to keep the cells growing, so the wan, and China are fast becoming work attracted little publicity. That major centers for human ES cell changed when two U.S. groups— research. Like their colleagues in Spotlight.Woo Suk Hwang (above) and Shin Young Moon grabbed acclaim one led by James Thomson of the the advanced scientific powers— for South Korea with their breakthrough work with ES cells. University of Wisconsin, Madi- including Japan, the United son, and another by John Gearhart States, and many European countries— [any field in] cutting-edge biology,” says of the School of researchers in the developing countries of Chunhua “Robert” Zhao, director of the Medicine in Baltimore, Maryland—almost Asia are racing to learn how to transform ES National Center for Stem Cell Research in simultaneously solved the problem of main- cells into human tissues and organs, which Beijing. “This could be our chance.” taining stable lines of ES cells in 1998 by could lead to treatments for conditions that Stem cell researcher George Q. Daley of growing them on “feeder” layers of mouse are now intractable, such as diabetes, Parkin- Harvard Medical School in Boston agrees: “I fibroblast cells. Bongso and colleagues from son’s disease, and spinal cord injuries. firmly believe they have an advantage.” Monash University in Melbourne, Australia, But there is one big difference: Unlike Although recent state funding initiatives in and Hebrew University in Jerusalem caught their colleagues in the United States and California and Wisconsin (see sidebar, up, creating their own stable human ES cell much of Europe, Asian scientists have the p. 662) should ease some of the constraints lines in 2000. full support of their governments. Because hobbling ES cell research in the United States, obtaining ES cells involves the destruction says Daley, such efforts are no substitute for Singapore of very early stage embryos, many Western federal support, which is still restricted. Singapore was quick to realize the scientific governments have placed heavy restrictions Asia does face challenges, however. and commercial payoffs of stem cell research. on the work. But across Asia, there is little These countries are still building their scien- “Given its huge potential, stem cell research of the conflict with prevailing religious and tific infrastructures, and many institutions has been identified as one of Singapore’s ethical beliefs that has Western countries must make do with older equipment. For niche areas,” explains Hwai Loong Kong, hesitating (see sidebar, p. 664). Govern- some groups, geographical isolation and lin- executive director of the Biomedical Research ments are ramping up funding for both gering language barriers hinder participation Council, a part of Singapore’s Agency for Sci- basic and applied stem cell work, setting up in conferences and complicate scientific ence, Technology, and Research (A*STAR).

new institutes, programs, and grant publishing. In China, a lack of coordination ES cells became a cornerstone of Singapore’s CREDIT: KIM KYUNG-HOON/REUTERS

660 4 FEBRUARY 2005 VOL 307 SCIENCE www.sciencemag.org State of the art. With a 25-person research team, Singapore’s ES Cell International is racing to create insulin-producing ES cells to treat diabetes.

$2 billion National Biomedical Science Strat- In March 2002, ES Cell recruited Alan stem cells into insulin-producing cells that egy, announced in June 2000 (Science, Coleman, former research director of PPL could be transplanted into patients with dia- 30 August 2002, p. 1470). Therapeutics in Edinburgh, U.K., and a betes. Robert Klupacs, ES Cell Inter- Kong says A*STAR is spending about member of the team that cloned Dolly the national’s CEO, says it hopes to start human $7.3 million per year to support stem cell sheep, to head its 25-person research team. clinical trials in 2006. Ronald McKay, a research, using both embryonic and adult The company is banking on its ability to turn stem cell researcher at NIH, says ES Cell lines, at the country’s national labs and International is definitely one of the teams through grants to university researchers. But Asia’s Stem Cell Firsts to watch, as is Singapore as a whole. that is only part of the story. Academic groups Although the company has yet to turn a also get funding from their universities and 1963 First cloned vertebrate (Asian carp) profit, concedes Klupacs, it has been able to the Ministry of Education. The amount can’t Tong Dizhou support its $6.1-million-a-year research pro- be pinned down, but the National University Institute of Oceanology, Chinese Academy of Sciences, gram with grants from Singapore, Australia, of Singapore reports that about a dozen Qingdao, Shandong Province, China and private investors. Science Bulletin (Chinese) groups are working on stem cells. Additional The U.S.-based Juvenile Diabetes money is coming through venture capital sup- 1973 First interspecies clone (European carp Research Foundation (JDRF) is also support- port for start-up companies working to com- DNA into Asian carp egg) ing stem cell research in Singapore. It pro- mercialize stem cell therapies and from for- Tong Dizhou vided a $600,000 grant to Bernat Soria of the eign funders attracted by Singapore’s wel- Institute of Oceanology, Chinese Academy of Sciences, University Miguel Hernandez de Elche in Qingdao, Shandong Province, China coming climate for ES cell research. ActaZoologica Sinica Alicante, Spain, to set up a lab in Singapore Among academics, Bongso continues to in 2002 to continue work he was prevented set the pace. In September 2002, he and his 1994 First isolation of human ES cells from doing in his native country. In February Australian and Israeli colleagues reported Ariff Bongso 2000, Soria reported that his group had dif- National University of Singapore the first propagation of human ES cells with- Human Reproduction ferentiated mouse ES cells into insulin- SCIENCE out using mouse feeder layers—a key producing cells that had alleviated dia- advance because the lines grown on mouse 2002 First propagation of betes symptoms in mice. He has been cells probably cannot be used for clinical human ES cells without use of extending that work to humans in his Sin- applications, given concerns about non- mouse feeder layers gapore lab. Although the Spanish govern- human pathogens. Bongso and his col- Ariff Bongso ment has since relaxed its restrictions, National University of Singapore leagues have turned over their cell lines and (plus colleagues from Monash Soria plans to keep a lab in Singapore. intellectual property to ES Cell International University in Melbourne, Australia, and “The Asia-Pacific is playing a very for commercialization. ES Cell owns six of Hebrew University in Jerusalem) important role in this research,” he says. the 22 human ES cell lines currently listed on Nature JDRF is also putting up half the cost of a the U.S. National Institutes of Health’s 2003 First isolation of embryonic stem $3 million fund—the other half is coming (NIH’s) Stem Cell Registry and has supplied (ES) cells from cloned human cells from A*STAR—to support other Singapore- more than 140 ES cell lines to researchers Woo SukHwang and Shin Yong Moon based stem cell researchers working in a around the world, second only to the Wiscon- Seoul National University number of fields, as part of a new, competi- Science

CREDITS (TOP TO BOTTOM):TO ES CELL INTERNATIONAL; CREDITS (TOP D. NORMILE/ sin Alumni Research Foundation. tively reviewed grant scheme. The founda-

www.sciencemag.org SCIENCE VOL 307 4 FEBRUARY 2005 661 N EWS F OCUS

U.S. States Offer Asia Stiff Competition new facilities and research support for scientists at the University of Wisconsin, Madison. Post–Proposition 71, a planned $375 million Proposition 71, the $3 billion initiative designed to catapult California public-private research institute, the Wisconsin Institute for Discov- into position as the world leader in research involving human embry- ery, has gained impetus. onic stem (ES) cells, is having a seismic effect across the United States. In New Jersey, acting Governor Richard Codey is pursuing a A few states—notably Wisconsin and New Jersey—are trying to regional approach. He has proposed allocating $150 million from become counterweights to California. Others are proposing more unspent bond income to construct the New Jersey Institute for Stem modest measures to make their states more attractive to stem cell Cell Research, a joint project of Rutgers University and the University researchers. Many legislators are trying to of Medicine and Dentistry of New Jersey. float initiatives despite substantial obstacles, Codey wants a ballot referendum next such as big budget deficits. But if they don’t November to raise $230 million to bankroll take action,“states that have made significant research grants over the next 10 years. investments in biomedical research”—Mary- In Illinois, members of the state Senate land and Massachusetts, to name two—”are failed narrowly in November to pass a bill genuinely concerned they are going to lose that would have allowed state funding for ES intellectual capital and resources,” says Daniel cell and nuclear transfer research. Now state Perry of the Coalition for the Advancement of Comptroller Daniel Hynes has designed a Medical Research in Washington, D.C. California copycat initiative: a statewide ref- Wisconsin—where the first human ES cell erendum in 2006 on a billion-dollar bond ini- line was derived in 1998—is moving deci- tiative. The Illinois Regenerative Medicine sively. The state is poised for a massive new Institute would be created from the sale of investment of $750 million in stem cell and States’ rights. New Jersey’s Richard Codey is $100 million in bonds per year for 10 other biomedical research over the next few one of several governors trying to lure stem cell years—repaid through a 6% tax on cosmetic years, including more than $500 million in research to his state. plastic surgery.

tion is investing in Singapore, says Chief Sci- ing on all types of stem cells in more than 30 vice director of rural and social develop- entific Officer Robert Goldstein, “because scientific teams across the country. Perhaps ment, says the guidelines are “aimed at push- there is excellent science, a good environ- 80 of them work with embryonic cells, a pro- ing forward our country’s stem cell and ther- ment, and really strong support for work that portion that is growing. apeutic cloning research.” can’t be done in [the public sector in] the As opposed to Singapore’s coordinated The competition among groups and the U.S.,” including deriving and working with national plan, China has a host of over- government’s reluctance to reveal informa- new stem cell lines. lapping initiatives from the central govern- tion make it difficult to judge China’s ment, cities and provinces, private enterprise, progress. But there is one sign of success: the China and even semiprivate venture capital funds growing number of foreign-trained Chinese Although numbers are hard to verify, China created by government agencies and the mil- scientists who are leaving comfortable posi- may be home to the largest stem cell program itary. Pei pegs the total 5-year research tions in Europe and the United States to work in Asia. The government does not release sta- budget at “more than” $24 million. (Dollars in their native land. Sheng Hui Zhen, an ES tistics, but Pei Xuetao of the National High go further there than in the West, given cell researcher at Second Medical Technology Research and Development Pro- China’s vastly lower labor and material costs University, says that “most major teams” in gram’s stem cell division estimates that and its allocation of almost 100% of funding the field now have U.S.- or Europe-trained China has “about 300 to 400” Ph.D.s work- to research, with little overhead.) scientists in senior positions. Perhaps more important than Sheng is a prominent example. She spent funding levels, says Xiangzhong 11 years at NIH before relocating to Shang- “Jerry” Yang of the University of hai in 1999, where she leads a 50- Connecticut, Storrs, is that China person team, funded mostly by the city. The has one of the most supportive group is attempting to create functioning environments for embryo research human ES cells by inserting the nucleus of anywhere in the world. Issued in adult human skin cells into rabbit eggs from December 2003, Chinese regula- which the nuclear DNA has been extracted. tions are quite “liberal,” says Yang. Sheng reported initial success in August There is a strict ban on human 2003 in Cell Research, a peer-reviewed jour- cloning for reproductive purposes. nal backed by the Chinese Academy of Sci- But for research cloning, the ences, but so far, no other lab has reported guidelines include little more than duplicating her work. a requirement that scientists com- The popular press described the work as a ply “with the principle of informed “cross-species clone,” sparking intense ethi- consent and informed choice” cal debate in the West. But Sheng dismisses when obtaining embryos from IVF talk of chimeric animals, noting that the only clinics or fetal tissue from aborted rabbit DNA in the cells is mitochondrial. Her fetuses. They also include a direc- goal, she says, is to design an alternative to tive for institutions to monitor human eggs for use in therapeutic cloning. Coming home. Deng Hongkui left his lab in New York for new compliance. Wang Yu, the Min- She suspects that when such work becomes

digs in Beijing’s Peking University. istry of Science and Technology’s feasible, the procurement of eggs, which are BOTTOM): PRESS;YANTO SHEN MIKE DERER/ASSOCIATED CREDITS (TOP

662 4 FEBRUARY 2005 VOL 307 SCIENCE www.sciencemag.org N EWS F OCUS

Other states are eyeing various strategies to beef up their stem Laws prohibiting nuclear transfer (therapeutic cloning) have been cell capacities. In Maryland, legislators are readying a proposal that passed in Michigan,Arkansas, Iowa, North Dakota, and South Dakota. would use tobacco-settlement money to open up $25 million annu- Missouri is contemplating one, although scientists are warning that ally for stem cell research starting in fiscal year 2007. Florida is poised the state will pay a price if it adopts such a ban.The Stowers Institute to become a major player now that the California-based Scripps for Medical Research in Kansas City, a major contributor to the bio- Research Institute plans to open its first branch in Palm Beach County. logical lifeblood of the state, has said it “would be forced” to build a And a private group, Cures for Florida, is campaigning for a $1-billion- planned second facility outside Missouri if the measure passes. plus state ballot initiative for ES cell research. Perry sees the state initiatives as evidence that the center of grav- Legislators in Massachusetts are chafing to get into the stem cell ity in research may be shifting away from the federal government. game, but because of the state’s large Catholic population, recent “After generations in which a single NIH” ruled the biomedical pro-research measures have been quashed by the legislature. But research roost,“it’s almost like the breakup of the Roman Empire.” Democrats, who are angling for the support of Republican Governor –CONSTANCE HOLDEN Mitt Romney, have vowed this year to push legislation to promote stem cell research through measures such as tax incentives.And in New York earlier this month, three legisla- tors proposed a 10-year, $1 billion bond ini- tiative that would finance the New York Stem Cell Research Institute. On the flip side, a number of states are attempting to close the door on research with human ES cells. Nebraska, South Dakota, and Louisiana have forbidden such research.

difficult and expensive to obtain, may be the weak link. “My Chinese lab does not have everything my NIH lab had,” says Sheng. “But here I can work on this important prob- lem, and there I couldn’t.” Some Chinese scientists have received backing for research that astonishes their for- mer Western colleagues. Trained in Min- nesota, Zhao of the National Center for Stem Cell Research in Beijing is working on stem cells from the bone marrow of aborted fetuses—work that cannot be done with fed- eral funding in the United States and that many states have banned outright. “We have the freedom to look at these problems from many angles,” he says.

South Korea and Taiwan To date in South Korea, the private sector has taken the lead in stem cell research. Three of ences, with a major focus on stem cells. working to derive their own mouse-feeder- the four groups that have established ES cell Hwang will be moving his lab to Bio-MAX. free cell lines and to learn to control differen- lines are at private IVF clinics, and for their Meanwhile, Moon continues to direct activ- tiation. Their first target, too, is insulin- breakthrough work, SNU’s Hwang and ities at the Korean Stem Cell Research Cen- producing cells. While ITRI focuses down- Moon relied on a culturing technique devel- ter. Established in 2002, the center has an stream, Academia Sinica, Taiwan’s premier oped at one of them. Figures for private sec- annual budget of $7.5 million to support collection of publicly funded science labs, is tor spending are not officially tallied, and 30 researchers. And last year, the govern- now ramping up a stem cell program focus- Hyun Soo Yoon, director of research at ment put $5 million into a new competitive ing on understanding basic stem cell biology. Seoul’s MizMedi Hospital, which has a team grant scheme for research related to thera- of 18 scientists and technicians working full- peutic cloning, stem cells, and xenotrans- Challenges time on stem cell research, also declined to plantation. Funding could rise to $25 mil- Although Asia’s stem cell efforts are coming disclose his group’s budget. lion per year by 2008. into their own, the region faces a number of Now the South Korean government In Taiwan, the government-affiliated challenges. Some worry that important stem wants to capitalize on the advances made by Industrial Technology Research Institute cell research is going unpublished because of Hwang and Moon. At Hwang’s home uni- (ITRI) is trying to nurture the island’s the intense interest in commercialization versity in Seoul, the government is spend- biotechnology industry by developing stem by Asian governments, companies, and ing $50 million over 5 years to set up the cell expertise. ITRI researchers were the first researchers. Unlike Western biotech compa- Bio-MAX Institute; its goal is to foster in Taiwan to start working with human ES nies, which often seek the limelight, repre- interdisciplinary research in the life sci- cells, in 2001. They have an 18-person group sentatives of private companies in both Tai-

www.sciencemag.org SCIENCE VOL 307 4 FEBRUARY 2005 663 N EWS F OCUS partly because his lab was preoccupied with Asian Countries Permit Research,With Safeguards the SARS emergency and partly, he says, because he wanted to secure worldwide Government officials, researchers, and ethicists in Asia readily link the region's general accept- intellectual property rights. ance of research using human embryonic stem (ES) cells to its dominant Buddhist and Confu- In China, researchers admit, the pen- cian religious-ethical traditions. But the countries of East Asia have also put a lot of thought, chant for secrecy is heightened by rivalry effort, and public debate into formulating policies that define researchers’ responsibilities, as and suspicion, which sometimes prevents well as oversight mechanisms to ensure that guidelines are followed. groups from sharing data, expertise, and Although broadly similar, the policies adopted throughout the region differ in details. China, equipment as freely as their colleagues in South Korea,Taiwan, and Singapore have all banned reproductive cloning with the intent of cre- the West. But they contend that this lack ating a child.All four regions also allow the derivation of ES cells from surplus in vitro fertilization of communication is exacerbated by Asian (IVF) embryos obtained with informed consent; China, in addition, allows researchers to use researchers’ continuing isolation from embryos from aborted fetuses or miscarriages. South the scientific mainstream. Korea's law stipulates that only embryos preserved for at Yang attributes some of this iso- least 5 years can be used. In each country except China, lation to what he calls Western bioethics advisory committees have proposed national researchers’ “inability to believe review boards to approve and oversee the derivation of new that top-rank research can come stem cell lines and each specific research project using them. from developing nations in Singapore and China allow the creation of embryos Asia.” The biggest challenge fac- through IVF for research purposes; South Korea and Tai- ing the region “is not the lack of wan forbid this. Countries are split on therapeutic financial resources or good cloning, or the use of adult somatic cells to create stem bench-level researchers but the cells genetically matched to the donor. Singapore and lack of leaders,” says Haifan Lin, China will allow it with the same oversight as for ES cells. a stem cell researcher at Duke South Korea has decided to restrict therapeutic cloning University in Durham, North to a limited number of groups and solely for work that Carolina, who serves on a grant can't be done using typical ES cells. The country's review committee at China’s national review board will decide which groups and proj- National Natural Science Foun- ects qualify. Taiwan's advisory committee "split 50-50” dation. All of these countries are on therapeutic cloning, says committee member Daniel trying to recruit researchers from Tsai, a physician on the faculty of National Taiwan Uni- outside their borders. Singapore versity. It put off a decision pending further study. has been the most aggressive, Singapore, South Korea, and Taiwan incorporated socie- partly because it is so under- tal views through high-level bioethics committees that staffed. Singapore also has held public hearings and made recommendations for the advantages in recruiting non- governments to codify into law. South Korea adopted a law Socially acceptable.Asian coun- natives, as English is the language of com- governing ES cell derivation and research in December tries are less encumbered by the merce and government and the city is rela- 2003. In September 2004, Singapore banned reproductive ethical dilemmas that have ham- tively cosmopolitan. “For me, it was Singa- cloning but left pending the creation of a national review strung research in the West. pore or nothing,” says ES Cell’s Colman. board. Under both laws, violators face prison sentences of Fifteen nations are represented on ES Cell’s up to 10 years or hefty fines or both. South Korea’s review board is now being formed. Singa- 25-person scientific team. pore’s and Taiwan’s need enabling legislation. For now, researchers using ES cells in Singapore Some countries are already following must report their activities to the Ministry of Health. Until Taiwan passes legislation, says Tsai, China’s lead and targeting expatriate sons institutions are trying to follow the recommendations of the bioethics committee; anyone vio- and daughters. At Taiwan’s Academia lating the administrative ban on human cloning could lose a license to practice medicine or be Sinica, most of the half-dozen Ph.D.-level forced out of an academic post. researchers in the new stem cell group are Serious debate in China on stem cell research ethics began only in late 2001, after a team Taiwanese or Chinese researchers returning led by Chen Xigu of Zhongshan Medical University in Guangzhou claimed it had cloned from stints in the United States, the United embryos by inserting a child’s DNA into an enucleated rabbit egg.Although the news was met Kingdom, or Australia. Group leader John with skepticism, and the team never published its results, the report set off a public storm. Gal- Yu is a case in point. The former director of vanized by the furor, Chinese bioethicists held several meetings in 2002 and 2003, submitting experimental hematology at Scripps the results to a newly formed interagency committee of the ministries of Health and Science Research Institute in La Jolla, California, and Technology. Issued in December 2003, the committee’s “ethical guiding principles” are Yu says he was lured back by the opportu- much less formal than other nations’ regulations—they are fewer than 500 words long and nity to get in on the ground floor of an excit- specify no penalties for violation.Although the regulations are intended to “give researchers a ing new effort and the chance to work in his lot of freedom,” the bottom line is clear, says Deng Hongkui of Peking University:“There will be native region. no reproductive cloning in China.” –D.N. AND C.C.M. Yu and other Asian scientists say they view these questions about leadership, openness, efficiency, and labor power as wan and South Korea were reluctant even to Deng Hongkui, a former New York Uni- hurdles, not barriers, and are determined to name their research topics to Science. Speak- versity researcher known for his work on overcome them. And they say their Western ing under condition of anonymity, two Chi- HIV, moved in 2001 to Peking University. colleagues should expect to see more nese researchers confessed they had not fully Deng readily concedes that he has delayed headline-grabbing research results come informed their granting agencies of what submitting his research on the mechanisms out of Asia in the next few years.

they were doing. of differentiation for publication for 2 years –DENNIS NORMILE AND CHARLES C.MANN CREDIT:A. LASLETT,A. FILIPCZYK,AND M. STEM CELL CENTRE AUSTRALIAN AND PERA/MONASH INSTITUTE OF MEDICAL RESEARCH

664 4 FEBRUARY 2005 VOL 307 SCIENCE www.sciencemag.org Developmental Biology

path of least resistance, without much active guidance. But the recent work paints a sub- The Unexpected Brains Behind tler picture, in which guidance molecules provide precise attractive and repulsive cues Blood Vessel Growth to specific growing vessels, notes Christer Betsholtz of the Karolinska Institute in Two of the hottest fields in developmental biology—neural guidance and angiogenesis— Stockholm, . Work by Betsholtz and are beginning to merge as scientists find that similar proteins control both processes his colleagues revealed some of the first evi- dence for that precision. They showed in Whether in San Francisco or Singapore, ing cancerous tumors or to help them regrow 2003 that specialized cells at the tip of devel- almost everyone knows what the colors on a after illness or injury—the nerve proteins oping blood vessels are attracted by slight traffic light mean. But how did red, green, offer a wealth of new targets to manipulate. changes in the concentration of VEGF. That and yellow get chosen? It turns out railroad reminded many biologists of what they see at signals were already using these colors to The tipping cell the front of extending axons: the long exten- guide trains. And the railroad industry may One of the first signs of flirtation between sions of a nerve cell that reach out and con- have gotten the idea from the electrical the two fields came in 1998: Michael Klags- nect with other cells. “There are certainly industry, which apparently used red to show brun of Children’s Hospital in Boston and some differences,” says Ruediger Klein of that a motor was stopped and green to signal his colleagues reported in Cell that neuro- the Max Planck Institute of Neurobiology in that it was running. When something works, pilin, a cell surface protein originally identi- Martinsried, Germany, “but if you look at the why not use it more than once? fied as a receptor for a signal that guides pictures [from Betsholtz], the tip cells look Evolution follows that principle too, as growing nerves, also responds to VEGF very much like an axon’s growth cone, researchers studying the growth of blood (Science, 27 March 1998, p. 2042). Klags- extending and sensing the environment and vessels and nervous systems are beginning responding to cues.” to appreciate. Scientists probing the devel- And the tip cells seem to respond to at opment of the veins, arteries, and capillaries least some of the same cues as growth cones. that guide nutrients and oxygen to cells are Last November in Nature, a group led by finding more and more evidence that the Anne Eichmann of the College of France in genes and proteins that were first discov- Paris described how those tips respond to ered to guide growing nerve cells also direct netrins, a family of secreted proteins that help blood vessels. attract some axons and repel others during the Decades of work by formation of the spinal cord. How nerve cells detailing the complex interactions of those react to netrins depends on which receptors cues is now giving researchers who study they express, and, the new work shows, blood angiogenesis—the growth of blood ves- vessels can also react in different ways to the sels—a boost in their understanding of the chemicals. Eichmann, with Peter Carmeliet vascular system. “Neurobiology has made an of the University of in Belgium and immeasurable contribution to angiogenesis,” Mark Tessier-Lavigne of says David Anderson of the California Insti- in California and their colleagues, reported of Technology in Pasadena, who noticed that the gene for one of the previously identi- some of the first overlaps. “All the insights fied netrin receptors, called Unc5b, is we’ve gained from studying these signaling expressed in the tip cells of developing blood systems in the nervous system have put us in vessels. When the team created mice and a much better position to understand how zebrafish that made a faulty version of they work in the vascular system.” UNC5B, the vascular system of the mutant At the same time, one of the most power- animals had far more sprouts and branches ful triggers of blood vessel growth, a protein than normal—suggesting that the tip cells called vascular endothelial growth factor were impervious to a “stay away” signal from (VEGF), is turning up in nerve cells and Looking for direction. The end of a developing netrins. Indeed, the team subsequently may play a key role in keeping them healthy blood vessel sends out sensory tentacles that showed that Netrin 1 causes the sprouts of rat and alive. “There is remarkable overlap in resemble the growth cones of axons. blood vessels growing in culture to retract. the use of these [signaling] systems,” says But that is not the whole story. In a paper David Ginty of Johns Hopkins University in brun’s observation “was an amazing discov- published nearly simultaneously in the Pro- Baltimore, Maryland. ery,” Ginty says, although in hindsight it ceedings of the of Sciences, These insights not only are inspiring a makes perfect sense, because both the blood Dean Li and his colleagues at the University of new respect for the complexity and precision vessel and nervous systems are “vast net- Utah, Salt Lake City, showed that Netrin 1 can of growing blood vessels, but they also have works of complicated connections.” Later also encourage the growth of new blood ves- potential medical implications. Animal trials that year, Anderson and his colleagues sels, suggesting that the molecule may reprise suggest that VEGF is a potential weapon reported that another set of neuronal guid- its sometimes attractive, sometimes repellent against amyotrophic lateral sclerosis (ALS), ance molecules, cell surface proteins called role in the vascular system. an incurable disease that attacks nerves and Ephrin B2 and EphB4, were also present in Eichmann and her colleagues have come gradually paralyzes its victims. And for the developing vascular system. across hints of other roles for netrins and those trying to control the growth of blood Before these new observations, blood their receptors in blood vessel development.

CREDIT: A.ABRAMSSON/KAROLINSKA INSTITUTE vessels—either to stop them from support- vessels were largely thought to form along a They found that the UNC5B receptor is

www.sciencemag.org SCIENCE VOL 307 4 FEBRUARY 2005 665 Grasp the Proteome™ Western Blotting Bad blot Restore™ Western Blot Erase-It ® Stripping Buffer Bad blot Background Eliminator whatchya’ gonna do?

Optimize experimental Remove background conditions without reblotting. from overexposed film.

Qentix™ Western Blot StartingBlock™ Signal Enhancer X Blocking Buffer Don’t let a bad blot happen to you. Achieve the highest signal:noise ratio Generate more signal from possible with these four new Western Prevent background the same bands. blotting products from Pierce. before it happens.

FREE Western Blotting Handbook! Log on to www.piercenet.com or call www.piercenet.com 800-874-3723 to request your FREE Western Blotting Handbook!

Tel: 815-968-0747 or 800-874-3723 • Fax: 815-968-7316 • Customer Assistance E-mail: [email protected] Outside the United States, visit our web site or call 815-968-0747 to locate your local Perbio Science branch office (below) or distributor Belgium & Dist.: China: France: Germany: : The Netherlands: United Kingdom: Switzerland: Tel +32 (0)53 83 44 04 Tel (8610)8048 9552 Tel 0800 50 82 15 Tel 0228 9125650 Tel 852 2753 0686 Tel 076 50 31 880 Tel 0800 252185 Tel 0800 56 31 40 [email protected] [email protected] [email protected] [email protected] [email protected] [email protected] [email protected] [email protected] © Pierce Biotechnology, Inc., 2005. Pierce products are supplied for laboratory or manufacturing applications only. Erase-It®, Qentix™, Restore™ and StartingBlock™ are trademarks of Pierce Biotechnology, Inc. N EWS F OCUS widely expressed in arteries, which deliver phorins keep developing blood vessels on altered versions of the protein. The mice oxygen-rich blood to tissues, but it is appar- the straight and narrow. According to their seemed to develop normally but became ill ently absent in veins, which return oxygen- research, zebrafish and mice lacking sema- as adults. “To our surprise, we found that depleted blood to the heart. phorins or their receptors develop strikingly they had motor neuron degeneration similar The early work on ephrin and Eph mole- disorganized vessels. to that seen in ALS,” says Carmeliet. cules from Anderson and his colleagues had A role in blood vessel growth for a fourth Normally, VEGF is expressed in showed a similar pattern. In their 1998 paper category of neuronal guidance molecules— response to low oxygen levels—it attracts that established some of the first links the Slit proteins and their Robo receptors— new blood vessels to tissues that are short of between neuronal guidance and angiogene- may be emerging as well. In 2003, Jian-Guo oxygen. Carmeliet’s mice carry a mutation sis, the team showed that Ephrin B2 is Geng of the Shanghai Institutes for Biologi- that prevents that oxygen-dependent increase in expression, suggesting that per- haps a lack of VEGF nerves vulnera- ble to hypoxia. Indeed, in studies of nerves in culture, introducing VEGF seemed to help the cells survive stressful conditions such as low oxygen or serum deprivations. There are early hints that VEGF might play a role in some human ALS cases as well. In a study of 2000 people in England and Sweden, Carmeliet and his colleagues found that those carrying a certain version of the VEGF gene, one which seems to lower its overall production, were 1.8 times more likely to develop ALS than the general population. Carmeliet and his colleagues have tested in animal models of ALS whether increas- ing production of VEGF combats the condi- tion. In one rodent trial, they injected into muscles a rabies virus, which homes in on and infects nerve cells, modified to churn out VEGF. The mice that received the virus took longer to develop ALS-like symptoms and survived longer than their untreated counterparts. Working with a rat model of ALS, the team has also injected the VEGF Follow me. In developing chick skin, arteries (red) align closely with nerves (green). protein directly into the cerebral fluid and documented similar benefits. The team is expressed in arteries but not veins. Con- cal Sciences at the Chinese Academy of Sci- now preparing human trials, Carmeliet says, versely, the ephrin receptor called EphB4 is ences in Shanghai and his colleagues which could be under way within 2 years. expressed in veins but not arteries. These reported that a wide variety of tumor cells Angiogenesis researchers are hoping data were the first sign that arteries and produce a protein called Slit2, and that that the molecules that originally held the veins are molecularly distinct at the earliest endothelial cells, the precursors of blood promise of regrowing severed or damaged stages of development. Klein and his group vessels, express the receptor Robo1. They nerves may pay off in another clinical area confirmed that finding several months later suspect that the tumor cells might be using as well: the fight against cancer. These and showed that the proteins could prompt Slit proteins to attract new blood vessels to researchers have been attempting to fight the growth of new capillaries. the growing tumor tissue. And in October, tumors by cutting off their blood supply— Anderson and his colleagues found Roy Bicknell of Oxford University and his essentially starving them to death. The find- another bond between developing nerves and colleagues reported evidence in the FASEB ing that neural guidance molecules influ- blood vessels. They showed that in the skin of Journal that a newly identified Robo recep- ence normal blood vessel growth has sug- developing chicks, arteries are guided in part tor, which they call Robo4, is present in gested a wealth of potential new targets, by the development of nerves, whereas veins areas where new blood vessels are forming. says Tessier-Lavigne: “There is every rea- are not. They also studied mice lacking Sem- son to believe [these molecules] will regu- aphorin3A, one of the proteins that neuro- Receptive nerves late pathological angiogenesis as well.” pilins recognize. These animals develop Neuroscientists are also learning from The discoveries in both fields may have badly misdirected nerves, and their develop- angiogenesis researchers. VEGF, the classic even wider impact. Eichmann and her col- ing arteries followed the deviant paths of the trigger of blood vessel growth, is showing leagues have shown that mice lacking nerves, providing more evidence that the sys- up more and more in studies of nerve growth neuropilin-2 have defects in their lymph tems are closely intertwined. and development. The first clues to VEGF’s systems. Similarly, in the 1 February issue That observation is consistent with work neuronal role came from experiments in of Genes and Development, Klein and his on semaphorins by two other groups, one led Carmeliet’s lab at the University of Leuven colleagues describe how mice lacking by Ginty and the other by Brant Weinstein of in Belgium. To sort out some of the multiple Ephrin B2 develop major defects in their the National Institute of Child Health and roles VEGF plays in vascular development, lymphatic systems as well. Nature, it seems, Human Development in Bethesda, Mary- Carmeliet and his colleagues created several has made the most of a good idea.

CREDIT:AND D.ANDERSON/CALIFORNIATECHNOLOGY Y. INSTITUTE OF MUKOYAMA land. Each showed last year that sema- strains of mutant mice that carried slightly –GRETCHEN VOGEL

www.sciencemag.org SCIENCE VOL 307 4 FEBRUARY 2005 667 668 C current funding policiesaredesignedto current their income.Some researcherssuspectthat as many studentsaspossible tomaintain for government researchmoney andattract which universities must viewitheachother mia isnow acompetitive marketplace in And theoncecozyworld ofBritish acade- ence, forensicscience,andmedia studies. sci- coursessuchassports career-specific asstudentsareluredintomore falling classes hasbeensteadilyand chemistry forces areatwork, too.Demandforphysics University’s SchoolofChemistry. Butother level,” says PhilipKocienski, headofLeeds tooperateatevendepartments aminimum science tion “hasnever beenable tofinance physics andchemistry. This public contribu- for expensive lab-basedcoursessuchas U.K. universities, isnotproviding enough ment, which isthemainsource ofmoney for and they generally respondthatthegovern- science. sports biosciences andonstrengthsinphysics and ment andconcentrateonanew schoolof depart- in Decembertoclosethechemistry Nevertheless, theuniversity’s senatevoted 21% riseinapplicationstostudychemistry. 2004–05 academicyear, hadseena university departments. And duringthe (RAE), agovernment schemethatgrades by the2001Research Assessment Exercise national standardofexcellence, asjudged not failing: Almost allitswork meta because Exeter’s was department chemistry sity. caseparticularly Itwas asurprising fromtheuniver- degree back anhonorary andKrotothreatenedtohand department, announced planstocloseitschemistry news latelastyear when Exeter University Kroto oftheUniversity ofSussex. disaster,” says NobelistHarry chemistry Physics hassuffered asimilardecline.“It’s a offer degrees. chemistry undergraduate now lessthanhalfofallU.K. universities closures,and steady streamofdepartment Over thepastdecade,they have announceda ties intheUnitedKingdomseemtothinkso. doesn’t orphysics? teachchemistry Universi- institution stillworthy ofthatmoniker ifit whole, theworld, ortheuniverse. Butisan from theLatin ..Universities U.K. changes in society—is decimating chemistry andphysicschanges insociety—is decimatingchemistry departments Britain’s tofavor scheme thehighest-scoring research teams—abettedby other Demise ofPhysics andChemistry andthe ‘Darwinian’Funding AMBRIDGE Ask researcherswhy thisishappening, closuresbecameheadline Department U.K.— , universitas— The word “university”— suggests the ERAY20 O 0 CEC www.sciencemag.org SCIENCE VOL 307 4 FEBRUARY 2005 All alone. increase thenumber ofstudentsgoinginto government hassuccessfully worked to stable, Prime Minister Tony Blair’s Labour numbers ofapplicationshave stayed fairly dents asthey oncewere. Although absolute popular amongprospective university stu- Physicalone criticalfact: sciencesarenotas No amountofnew money would getaround Get ’emwhile they’re young tain: Nonew money willbeavailable. nobodyknows.departments Onethingiscer- will halttheclosureofphysical science technology, andmathematics. Whether this which includesallofscience, engineering, strategic areasof study,protect five oneof for England(HEFCE)tolookintoways to asked theHigherEducationFundingCouncil CharlesClarke then–Education Secretary ment torethinkitsneutrality. InDecember, drawal hasforcedthegovern- fromchemistry autonomy. Buttherow over Exeter’s with- approach sofar, respectingtheuniversities’ says Kocienski. “It’sdepartments. aDarwinianexercise,” resources inasmallernumberofsuper- weed outtheweak andconcentrate The government hastaken ahands-off Fewer and fewer U.K. high school studentswant highschool degrees. chemistry Fewer andfewer U.K. degrees inthosesubjects.JohnEnderby,degrees physics by orchemistry teacherswith As aresult,few highschoolpupilsaretaught foradvancement.salaries andopportunities teaching; othercareersoffer muchbetter into problem attractingsciencegraduates room. Britain’s schoolsystemhaslonghada on thelackofgoodrolemodelsinclass- but many blame science’s decliningappeal course over anotherisnotasimplequestion, University ofLondon. gist GraemeDavies, vicechancellorofthe serious supply-side problem,” says metallur- chosen majorfromthebeginning.) “Thereisa ticular subject,andthey specialize intheir students apply touniversities tostudyapar- smaller andsmaller. (IntheUnitedKingdom, fraction goingintophysical sciencesgrew higher education. As thetotalexpanded, the Science and Technology Committee,says Labour’s chair oftheHouseCommons anything togetstudents.” Gibson,now says university administrators “willteach the University ofEast Anglia inNorwich, headofbiologicalson, former sciencesat that career. MemberofParliament IanGib- courses thatappeartoprovide trackto afast seem glamorous,anduniversities now offer sion hasmadejobsinforensics,forexample, studying abasicscience.Meanwhile, televi- Few of highschoolstudentsseethebenefit cause: We don’t value teachers.” ments is“asymptomoftheunderlying London, says thecrisisinsciencedepart- president oftheInstitutePhysics (IOP) in What motivates teenagerstochooseone Other socialincentives areatwork, too.

CREDIT: GETTY IMAGES N EWS F OCUS

police chiefs have told his committee that they don’t want such graduates. What they Physical Sciences’ Declining Popularity need are “good and physicists.” , president of the Royal 2.0

Society of Chemistry, says “it is up to us” to Chemistry Number of institutions offering make careers in science attractive. Physics and astronomy 100 courses (dotted line) Follow the money Attracting students is not enough to keep a 1.5 department afloat, however, as Exeter’s expe- 80 rience has shown. Many believe that govern- ment funding policies are quietly changing 60 the shape of higher education by channeling research funding into science powerhouses 1.0 while leaving other departments to founder. 40 Government funding to universities is dis- tributed by HEFCE and partner councils in 20 Scotland, Wales, and Northern Ireland. For the current academic year, HEFCE, by far the 0.5 0 Percentage of students enrolled in courses of students enrolled Percentage largest of the four councils, will distribute 1995 1997 1999 2001 2003 $11 billion to English higher education insti- tutions, of which $7.2 billion goes in support of teaching and $2 billion for indirect costs research output. In 1992 HEFCE launched the to think like business people and make deci- associated with research. The teaching por- RAE, its quality-control survey, which it sions on purely financial grounds. “There is a tion is divvied up according to how many stu- repeats roughly every 6 years. Specialists in lack of understanding among academic big- dents the university enrolls and how expen- each subject rate the research in all university wigs about the needs of chemistry and sive their courses are to teach. So each research departments and grade them on a scale physics,” he says. Kocienski, voicing a pes- humanities student earns a university $6600, from 1 (the lowest) to 5*, the score reserved for simistic view, says the current total of about while each undergraduate in lab-intensive departments with “international excellence” in 40 chemistry departments may dwindle fur- subjects such as physics and chemistry, for more than half of the work submitted for ther to just 20: “I suspect that the government example, wins the university 1.7 times as review. These grades have a major impact on has this number in mind, too.” The physics much ($11,000). Medics, dentists, and vets funding, so before each new RAE, departments community is concerned that as closures con- earn a fourfold boost ($26,000). scramble to hire the hottest new researchers in tinue, ever-larger swaths of the country will Many researchers argue that this extra the hope of bumping up their rating. be left without any physics department. Stu- funding is not enough to cover the costs of Most U.K. research departments cluster dents may have to travel farther from home to lab buildings, materials, and support staff. around the top end of the scale, with the peak study physics, the IOP warns, and businesses “Chemistry is expensive to teach,” says of the curve around the boundary between will not be able to work with local researchers Campbell, and HEFCE provides “woefully grades 4 and 5. But following the 2001 RAE, on R&D projects. inadequate funding.” Enderby agrees: “In all many departments were shocked when the In Scotland, universities are already try- subjects the full cost of teaching is not met, government decided to focus on the top ing to counter the trend by taking a prag- but the shortfall is greatest for the laboratory achievers, pushing more of HEFCE’s matic approach: They are teaming science sciences.” HEFCE spokesperson Philip research funding into the highest-rated departments together for greater strength Walker counters that the allowances are departments. Since that assessment, depart- rather than letting the weakest go to the wall. based on a study of what universities actu- ments rated lower than 4 have received no Six Scottish physics departments have ally spend. “We have to have a fair and trans- research funding from HEFCE; those rated 5 formed SUPA, the Scottish Universities parent means to allocate the money,” he and 5* get approximately three times as much Physics Alliance, and chemists from four says. In any event, Walker points out, once per researcher as those rated 4. And since universities will meld into two superdepart- HEFCE has done its calculations, the money 2001, many 4-rated departments, such as ments: EastCHEM and WestCHEM. Each is given to the university as a lump sum. chemistry at Exeter, have found themselves of these bodies will be entered into the RAE “Universities can allocate the money inter- fighting for survival. as a single department. Last November, nally as they want.” The implication is that Although the RAE is a painful process, it’s these initiatives won $70 million for the next Exeter itself bears the chief responsibility widely credited with having improved the 4 years from Scottish funding bodies. “We for the choices it made. “Exeter’s chemistry quality of research in the United Kingdom. have a vision of where we are going in chem- department was not a dying animal,” says But many think it may have gone too far, and istry and physics,” says Edinburgh’s Chap- Stephen Chapman, head of the School of HEFCE is reviewing the system before the man. “We’re not going to close things Chemistry at Edinburgh University. “It was next RAE in 2008. “The RAE aims to starve because one department is not doing well.” shot rather than left to die.” out the weak, and it’s been quite effective. But Other changes may be coming. Education now it’s cutting into flesh rather than fat,” says Secretary Clarke’s decision to consider pro- Academic cattle market Kocienski. “Vice chancellors are all too ready tecting strategic subjects is a sign that the gov- Departments that find they cannot manage to use the RAE to cull expensive depart- ernment may have concluded that it cannot with the teaching grant from HEFCE often end ments,” adds Kroto. govern higher education by a form of natural up subsidizing teaching from their research selection. “Do we need every department to be income. But not all departments have this lux- Cooperation not competition? world beating?” asks Enderby: “No. Do we ury, as HEFCE research grants vary greatly in Gibson thinks the current crisis is the result of need a widespread education in physics? Yes.”

SOURCES: ; SOCIETY OF CHEMISTRY ROYAL size depending on the quality of a department’s politicians forcing university administrators –DANIEL CLERY

www.sciencemag.org SCIENCE VOL 307 4 FEBRUARY 2005 669 RANDOM SAMPLES Edited by Jennifer Couzin

After the Earth Moved NEW DELHI—Indian geologists are scrambling to remap the Andaman and Nicobar islands after the devastating earthquake and tsunami that struck South Asia on 26 December. Prithvish Nag, India’s surveyor general, said at a scientific meeting here last month that the 700-kilometer island chain in the Bay of Bengal has moved southeast toward Sumatra by as much as 1.25 meters and has been twisted in a counterclockwise direction. At the same time,the combination of sinking land and rising water levels has meant that the sea has swallowed roughly 1 vertical meter of coastline. India plans to spend at least $25 million to document the island’s new geomorphol- ogy,Nag said.The dozen or so Global Positioning System control locations on the islands needed to be recalibrated after having been thrown for a loop by the magnitude 9 earthquake. Nag said the remapping must be done quickly so that the government can provide advice on where to relocate residents now temporarily housed in refugee camps on the islands, which support a large air force base and a substantial navy presence.

deliveries and unintended pregnancies, A Nose The Burden of Sex which cause psychological distress, as Sex has its repercussions, especially in the causes of disability. for United States.There, premature death and They found that U.S. women Survival disability linked to sexual behavior is triple bear more of the brunt than men. that in other wealthy countries, researchers In particular, they suffer from curable Encountering have found. infections, such as chlamydia, and their a robber crab Previously, epidemiologists have consequences, particularly infertility, might send counted cases of sexually transmitted the team reports in the February issue you running diseases (STDs) and deaths caused by of Sexually Transmitted Infections. in the opposite them. But that’s only part of the picture, For men, HIV is by far the leading direction.The says Shahul Ebrahim of the Centers for problem.The heavy toll is not surprising, world’s largest Disease Control and Prevention in given the higher incidence of HIV and land-living arthropod weighs up to 4 Atlanta, Georgia. Some STDs boost the unintended pregnancies in the U.S., says kilograms and steals anything it can nab risk of a second infection or cervical epidemiologist Ward Cates, president of with its formidable pincers. It turns out cancer or cause infertility. Ebrahim Family Health International, a nonprofit the crab has another unusual feature: and colleagues also included premature in Research Triange Park, North Carolina. It has evolved to smell on land much the way insects do.The finding is “cool,” says Leslie Vosshall, an olfactory researcher at Da Vinci Rockefeller University in New York City. Bill Hansson, a chemical ecologist at the Swedish University of Agricultural

Discovery CODEX ATLANTICUS Sciences in Alnarp, Sweden, measured the Five small rooms wedged electrical activity of olfactory neurons in between a Florentine military the crab’s short antennae while puffing institute and a monastery scents over these sensory organs.The were apparently once a electrical activity spurred by certain odors canvas for Leonardo Da Vinci. was the same in robber crabs as in insects. Last month,three researchers And like insects, the crabs weren’t drawn at the Italian Institute of Military Geography announced that they’d stumbled upon to just any smell; they favored chemicals, mural paintings after an unused staircase was demolished.The soaring birds (see picture) like dimethyltrisulfide, that are released closely resemble drawings by the 16th century master (inset).“To us, there seems to from decaying meat. be no doubt,” says Roberto Manescalchi, one of the discoverers. Superimposing images Hansson’s team also found that the short of the wall paintings on drawings in Leonardo’s Codex Atlanticus, he says, revealed a antennae are structurally similar to those perfect match. of insects and have a wrinkly lining, which Still, questions remain: Several experts have since visited the rooms, but they can’t probably helps sense odors.This is a “great resolve whether the murals are by Leonardo himself or one of his pupils, says Cristina example of convergent evolution,” says Acidini, head of an institute for the restoration of works of art in Florence operated by Hansson.The researchers report their results Italy’s Ministry of Culture. in the 26 January issue of Current Biology. CREDITS (TOP TO BOTTOM): IMAGING;TO SPACE AND BILL S.CREDITS (TOP MARCUS STENSMYR HANSSON;

670 4 FEBRUARY 2005 VOL 307 SCIENCE www.sciencemag.org RANDOM SAMPLES PEOPLE Edited by Yudhijit Bhattacharjee

JOBS TWO CULTURES involves predicting bank robbers’ Moving on. In a different political climate, chemistry Nobelist Math is hip. After 37 years at behavior: where and when they says he might have been headed for the National the California Institute of Tech- are going to hit next.” Institutes of Health. Instead, the outspoken Agre—a vigorous nology, mathematician Gary Lorden is supposed to lend supporter of Democrat John Kerry in last fall’s presidential Lorden finally authenticity to campaign—is heading to Durham, North Carolina, to become has students the mathematics Duke University Medical Center’s vice chancellor for science asking him for on the show, and technology, a newly created position that combines advis- his autograph. which he’s done ing with advocacy. But it’s a televi- so far by suggest- A at Johns Hopkins Uni- sion show, not ing changes to versity in Baltimore, Maryland, since his lectures, that the dialogue and 1981, Agre says he was one of three has made him a having graduate candidates under consideration to be celebrity. students write director of the National Heart, Lung, The chair of equations on and Blood Institute (NHLBI) before Caltech’s math Charlie’s dropping out last summer because he department, black- couldn’t stomach the Bush Adminis- Lorden (inset) boards. tration’s stance on key science issues is an adviser Lorden such as stem cell research. Last week to Numb3rs,a hopes that Elizabeth Nabel accepted the position cop drama on Numb3rs at NHLBI (Science, 28 January, p. 495). CBS featuring will help But Agre insists he is “not going to mathematician students Duke in any partisan way.” Instead, Charlie (David Krumholtz), realize that mathematics isn’t he’ll help set research priorities for who helps his detective brother just for fusty academics. the medical center, serve as the university’s spokesperson on key solve crimes.“It’s kind of like science policy issues, and act as a “cheerleader for science” by Sherlock Holmes on steroids, AWARDS promoting science literacy among the general public. “I have a where the steroids are math- weakness for championing causes,” he says. ERGENCY MEDICINE; KEVIN STEARNS/ ematics and computer science,” Academy honors. A veteran of says Lorden.“In one episode, the campaign to wipe out they’re tracing the outbreak smallpox has been awarded the cation in Africa Crafoord prize. A transatlantic of a disease from a kind of National Academy of Sciences’ in the 1960s led trio of cosmologists has won terrorist attack. Another one (NAS’s) highest honor, the Pub- to the successful the 2005 Crafoord Prize, lic Welfare Medal. “ring vaccination” awarded by the Royal Swedish Got any tips for this page? E-mail Epidemiologist William H. strategy of inoc- Academy of Sciences. [email protected] Foege’s work on smallpox eradi- ulating close ’s James contacts of Gunn and James Peebles and PIONEERS infected people. Cambridge University’s Martin A flood of interest. Since the tsunami devastated South Asia, Foege later directed the Centers Rees will share the $500,000 Evan Variano, a graduate student in civil and environmental for Disease Control and Preven- prize for work on how the engineering at Cornell University,has taught more than 500 middle tion, steered the international universe evolved from a smooth school and high school students how tsunamis develop and travel. Task Force for Child Survival and primordial soup of particles Variano sets up a narrow, water-filled tube with a balloon inside, Development, and worked on and radiation into the present then inflates the balloon.The result is a sudden displacement of eradicating Guinea worm dis- cacophony of galaxies and clus- water,sending tsunami-like waves to the far end,where the amount ease and river blindness at the ters.All three are theorists, but of flooding Carter Center. In 1999 he joined Gunn is also lead project scien- depends on the Bill and Melinda Gates tist of the Sloan Digital Sky Sur- the shape of Foundation, where he helped vey, the largest project to date a miniature start programs including hepati- to map the three-dimensional coastline. It tis B immunization and AIDS distribution of galaxies. works better vaccine development. His work “These are all very large than a lecture, “has changed the world as we names in cosmology,” says he says: “The know it,” says NAS home secre- theoretical astrophysicist demo catches tary John Brauman. Vincent Icke of Leiden Univer- their atten- Now retired, Foege will be sity in the Netherlands. tion, and they honored 2 May along with Their career contributions, are curious to 17 other winners of various he explains, have been “a real know more.” academy awards. boon to the field.” CREDITS (TOP TO BOTTOM):TO PICTURES CORP.; PARAMOUNT CREDITS (TOP (INSET), BOB PAZ/CALTECH; DUKE UNIVERSITY MEDICAL CENTER;ACADEMIC EM SOCIETY FOR

www.sciencemag.org SCIENCE VOL 307 4 FEBRUARY 2005 671 Announcing 2005 PNAS chemistry special feature issues

Chemical theory and computation Long range electron transfer Cluster dynamics and chemistry Molecular electronics Intermolecular structure and dynamics Surface chemistry

PNAS encourages submissions of manuscripts in the chemical sciences

Contact Dr. Sarah Tegen at [email protected]

PNAS 2003 impact factor 10.2

Rapid time from submission to publication

All content freely available after 6 months

www.pnas.org

Proceedings of the National Academy of Sciences of the United States of America www.sciencedigital.org/subscribe For just US$130, you can join A A A S TODAY and star t receiving Science Digital Edition immediately! www.sciencedigital.org/subscribe For just US$130, you can join A A A S TODAY and star t receiving Science Digital Edition immediately! Letters to the Editor Letters (~300 words) discuss material published LETTERS in Science in the previous 6 months or issues of general interest. They can be submitted through the Web (www.submit2science.org) or tribes) (4); the abacus (Chinese) (5); populations revealed strongly density- by regular mail (1200 New York Ave., NW, Roman and Arabic numerals; and so forth. dependent population change and hence a Washington, DC 20005, USA). Letters are not The systems are all combinatorial: The use carrying capacity. acknowledged upon receipt, nor are authors of a base (e.g., 5, 10, 20) permits forming The suggestion that multispecies systems generally consulted before publication. new numbers by combining existing num- are unlikely to show density dependence is Whether published in full or in part, letters are bers. They are also recursive: New numbers erroneous. In contrast, evidence is emerging of subject to editing for clarity and space. are formed by adding one. Both recursion the very widespread occurrence of density and a combinatorial approach are long- dependence (3), even in complex marine sys- standing human capacities evident in, for tems (4, 5). Detection of such effects typically Evolution Versus example, both language and (to a lesser takes over four generations (6); well over a Invention extent) social behavior. hundred years might be needed to detect den- Conceivably, the same contingencies led sity dependence in an elephant population. THE PAPERS BY R. GELMAN AND C.R.GALLISTEL both to the invention of written language (“Language and the origin of numerical and to exact numeracy. Writing arose as a concepts,” Viewpoint, p. 441), P. Pica et al. belated consequence of the transition from (“Exact and approximate arithmetic in an foraging to agriculture. Farming resulted in Amazonian indigene group,” Report, p. surplus goods, necessitating a system for 499), and P. Gordon (“Numerical cognition marking the goods, identifying ownership without words: evidence from Amazonia,” of casks of oil and the like [(1), p. 8]. Report, p. 496) in the Special Issue on Although hunter-gatherers had little Cognition and Behavior (15 Oct. 2004), need for exact numbers, one can imagine taken jointly, establish two key points: that no owner of stored goods would wish to Animals as well as humans have an analog receive three casks of oil when he had system for representing numeracy. The ana- stored four, nor would the individual who log system, although yielding only approx- had stored the goods wish to return five imate answers, supports complex numeri- casks when he had stored four. cal computations including comparison, DAVID PREMACK* AND ANN PREMACK addition, and subtraction. Somis, CA, USA. E-mail: dpremack@aol .com Hunter-gatherer groups, whose lan- *Emeritus Professor of Psychology, University of guages contain only a few number words Pennsylvania (“one, two, many” in one case; “one” to References The application of nonequilibrium “five” in another), pose a problem. Despite 1. D. Premack, A. Premack, Original Intelligence hypotheses to savannah has been chal- (McGraw-Hill, New York, 2003). having number words, they perform only 2. M. D. Coe, Breaking the Maya Code (Thames & Hudson, lenged on theoretical and empirical approximate numerical calculations, even London, 1999). grounds. Models indicate that in when problems contain only numbers for 3. M. Ascher, R. Ascher, Code of the Quipi: A Study in semi-arid areas are in long-term equilib- Media, Mathematics, and Culture (Univ. of Michigan, which they have words. Ann Arbor, MI, 1981). rium with a subset of their resources (7). Despite their interest and clarity, we sug- 4. P. Gerdes, Historia Math. 21, 345 (1994). Competitive regulation is now very clear in gest that the above-mentioned papers over- 5. Y. Li, S. Ran Du, Chinese Mathematics, a Concise a number of species of large herbivores in look two key points. First, hunter-gatherer History, translated by J. N. Grossly, A. W. C. Lun Africa, including wildebeest (8). A review (Clarendon Press, Oxford, UK, 1987). number words are not comparable to our (9) concluded there was no evidence of a own. Even the smattering of data reported paradigm shift to a nonequilibrium per- by these authors indicates that hunter- spective among those researching grass- gatherer number words name approximate Elephants, Ecology, and lands. Large species are unlikely to exhibit magnitudes, not exact sets of objects as Nonequilibrium? metapopulation dynamics (10). ours do. There is then no paradox in the fact The harvesting of elephants is, rightly, con- that hunter-gatherers perform only approxi- ELEPHANTS AND THRIPS MAY HAVE SOMETHING troversial. We suggest that the nonequilibrium mate numerical calculations. in common: It has been proposed that ele- perspective is unlikely to clarify how their pop- Second, it is not language per se that is phants in Africa do not reach carrying ulations might respond to management. critical for understanding the transition capacity because they inhabit “nonequilib- CLIVE HAMBLER,PETER A. HENDERSON, from analog to digital numeracy, but the rium” ecosystems with highly variable rain- MARTIN R. SPEIGHT change from foraging that did not require fall (“Space—the final frontier for econo- Department of Zoology, , exact numbers to technologies that did. mists and elephants,” E. Bulte et al., South Parks Road, Oxford OX1 3PS, UK. Humans, as hunter-gatherers, evolved a Perspectives, 15 Oct., p. 420). Similarly, it References system for analog numeracy [(1), p. 29]. has been proposed that thrips in Australia 1. H. G. Andrewartha, L. C. Birch, The Distribution and When forced to adopt new technologies— do not reach a carrying capacity because of Abundance of Animals (Univ. of Chicago Press, pastoralism, gardening, trade or barter, and climatic fluctuations (1). The nonequilib- Chicago, IL, 1954). 2. F. E. Smith, Ecology 42, 403 (1961). SCIENCE ultimately full-time farming—they then rium (density-independent) ideas of the 3. R. Lande et al., Am. Nat. 159, 321 (2002). invented systems for representing exact 1950s are being reworked as “state-of-the- 4. M. S.Webster, Ecology 85, 986 (2004). numeracy: dots, bars, and a shell standing art” ecological theory by Bulte et al. We 5. J. S. Shima, Oecologia 126, 58 (2001). 6. P. Rothery, in Insect Populations in Theory and Practice, for zero (Mayan) (2); ropes and knots should remember, however, that a more P. J. Dempster, I. F. G. McLean, Eds. (Kluwer Academic,

CREDIT: SMITH/ ORLA (Incas) (3); fingers and fists (sub-Saharan sophisticated analysis (2) of the same thrip Dordrecht, 1998), pp. 97–133.

www.sciencemag.org SCIENCE VOL 307 4 FEBRUARY 2005 673 L ETTERS

7. A.W. Illius,T. G. O’Connor, Oikos 89, 283 (2000). species but differ with their interpretation only are theoretically rigorous but also 8. A. R. E. Sinclair, C. J. Krebs, in Wildlife Population Growth of the term “nonequilibrium.” A more incorporate appropriate spatial and tempo- Rates, R. M. Sibly, J. Hone, T. H. Clutton-Brock, Eds. (Cambridge Univ. Press, Cambridge, 2003), pp. 127–147. important point, however, is that the sim- ral complexities of changing environments. 9. D. D. Briske, S. D. Fuhlendorf, F. E. Smeins, J.Appl. Ecol. 40, plistic bioeconomic models that we cri- LINDSEY GILLSON,1 KEITH LINDSAY,2 610 (2003). tiqued in our Perspective do not acknowl- ERWIN H. BULTE,3 RICHARD DAMIANA4 10. S. Harrison, in Large-Scale Ecology and Conservation 1 Biology, P. J. Edwards, R. M. May , N. R.Webb, Eds. (Blackwell edge the many other, complex interactions, Environmental Change Institute, Biodiversity Scientific Publications, Oxford, UK, 1994), pp. 111–128. apart from density dependence, that can Research Group, c/o Department of Zoology, also influence population size, nor the fact University of Oxford, Oxford OX1 3PS, UK. IT ISA PLEASURETO SEE ECONOMISTS BORROWING that density-dependent effects are only 2Amboseli Elephant Research Project, Amboseli from ecology (“Space—the final frontier for dominant at certain spatial scales (1). Trust for Elephants, Post Office Box 15135, Langata economists and elephants,” E. Bulte et al., We used the term “nonequilibrium” in 00519, Nairobi, Kenya. 3Department of Economics, Perspectives, 15 Oct., p. 420) because the its broadest sense to mean “not-at-equilib- Tilburg University, Post Office Box 90153, 5000 LE traffic has usually been in the other direction. rium,” rather than to imply that equilibrat- Tilburg, The Netherlands. 4School of Economics, But Bulte et al. are surely throwing the ele- ing, density-dependent processes are not University of Adelaide,Adelaide 5005,Australia. phant out with the bathwater when they important. It is well recognized in the eco- References assert that equilibrium ideas in population logical literature that environmental sto- 1. D. L. DeAngelis, J. C. Waterhouse, Ecol. Monogr. 57,1 (1987). ecology are often false. We know that tempo- chasticity can affect population size (1), but 2. R. H. Behnke Jr., I. Scoones, C. Kerven, Range Ecology at ral environmental variation prevents plant- it is only a very small subset of nonequilib- Disequilibrium: New Models of Natural Variability and systems from reaching a stable rium theories—those termed “disequilib- Pastoral Adaptation in African Savannas (Overseas Development Institute, London, 1993). equilibrium, but we also know what to do rium”—that assert that environmental vari- 3. J.Wu, O. L. Loucks, Q. Rev. Biol. 70, 439 (1995). about it, in theory, if our aim is to harvest ability can completely override the effects 4. S. Levin, Ecology 73, 1943 (1992). such populations sustainably (1). of biotic interactions. Rangeland ecologists, 5. J.Wu, J. L. David, Ecol. Model. 153, 7 (2002). “Nonequilibrium,” on the other hand, is a for example, have argued that herbivore shibboleth often used to invoke some myste- population density is largely determined by rious dynamical regime in which herbivores extreme events like drought, which in semi- are somehow not coupled to the dynamics of arid savannas are frequent enough to keep National Environmental their resources (2). The reality is simpler and populations at levels that are too low for Policy Act at 35 more conventional than that. Spatial environ- density-dependent effects like competition mental variation results in a patchwork of to operate (2). 1 JANUARY 2005 WAS THE 35TH ANNIVERSARY resources that vary in quality and accessibil- This disequilibrium viewpoint is of the signing into law of the U.S. National ity. A subset, the key resources (3), is what extreme, however, and most authors now Environmental Policy Act (NEPA). It has the herbivore population depends on to get agree that both density-dependent and envi- since been copied and enacted in many through the dry season, when mortality ronmental variables affect population size local U.S. jurisdictions and around the threatens. Modeling shows that, despite (3). Although density-dependent effects world, so we believe this is the proper time short-term fluctuation due to temporal sto- might cause populations to tend toward an to list some of its accomplishments and chasticity, the herbivore population is in equilibrium, it is likely that the position of continuing problems. long-term equilibrium with these key the equilibrium will change over time and First, the existence of the statute and its resources. In other words, herbivore popula- that factors such as rainfall, fire, disease, or implementing regulations have required tion size depends largely on the environ- human influence deflect populations away U.S. agencies to at least acknowledge that ment’s endowment of key resources and from a possible equilibrium or “carrying there are environmental consequences of hardly at all on the remaining parts of the capacity.” Rather, population size may be their actions. Second, the existence and habitat (4). Space may be the final frontier, envisaged as varying around an equilibrium publication of NEPA Environmental but the challenge is to identify and manage point (perhaps more usefully termed an Analyses (EA) or Environmental Impact the key resource, or equilibrial, parts of the “attractor”) and can move between two or Statements (EIS) have provided for much system, rather than to worry about the irrele- more domains of attraction. more public input into decision-making. vant nonequilibrium remainder. By charac- The debate in nonequilibrium ecology has But NEPA was designed to do more. It terizing the entire system as nonequilibrium, thus moved beyond the question of whether was meant to force agencies to “insure the and failing to make the distinction between density-dependent or environmental variables integrated use of the natural and social sci- types of resources, the nonequilibrium para- are most important. Since the early 1990s, dis- ences… in planning and decision-making.” digm suffers from the problem of not seeing cussion has focused on identifying the scales at The U.S. Council on Environmental the trees for the wood. which different processes predominate, and Quality (CEQ) regulations for NEPA ANDREW W. ILLIUS how to combine this knowledge in ways that implementation say that “NEPA’s purpose Institute of Evolutionary Biology, School of are ecologically meaningful (4). Recent is not to generate paperwork—even excel- Biological Sciences, , advances in hierarchical modeling, for exam- lent paperwork—but to foster excellent Edinburgh EH9 3JR, UK. ple, integrate nested processes in a spatially action,” and to lead agencies toward References explicit, hierarchical framework (5). Advances “actions that protect, restore, and enhance 1. R. Lande et al., Ecology 78, 1341 (1997). like these may provide the basis for spatially the environment” (1). That goal is fre- 2. A.W. Illius,T. G. O’Connor, Ecol.Applic. 9, 798 (1999). 3. I. Scoones, J.Arid Envir. 9, 221 (1995). defined—and more ecologically realistic— quently lost in the admittedly difficult 4. A.W. Illius,T. G. O’Connor, Oikos 89, 283 (2000). bioeconomic models in the future. process of producing an EA or EIS. In the meantime, equilibrium models In 1996, the average EIS was 570 pages, with density-dependent factors can provide although the CEQ regulations state it should Response an approximation of reality at certain be only 150 pages, or 300 for complex proj- WE AGREE WITH ILLIUS AND HAMBLER ET AL. (species specific) spatial scales. The chal- ects (1, 2). This mass of paper is frequently that density-dependent effects are impor- lenge is for economists and ecologists to unnecessary, as it is no more than the tant in the population dynamics of many work together to develop models that not description of the existing environment (3).

674 4 FEBRUARY 2005 VOL 307 SCIENCE www.sciencemag.org L ETTERS There are an enormous amount of envi- the process, but there are still aspects that Center for Science and Society, University of California ronmental data already gathered on almost need research and implementation if NEPA at Irvine, Irvine, CA 92697, USA. 5Environmental Law all of the United States. If these data were is to achieve its objectives. Program, Sierra Club, 85 Second Street, Second Floor, collected in one or more depositories, they DANIEL A. BRONSTEIN,1 DINAH BAER,2 HOBSON San Francisco, CA 94105, USA. could be reused and reduce the time and BRYAN,3 JOSEPH F. C. DIMENTO,4 SANJAY NARAYAN5 References 1 effort devoted to writing EISs (4) and could CARRS, Michigan State University, East Lansing, MI 1. 50 C.F.R. § 1500.1. serve as a quality control mechanism for 48864–1222, USA. 2Council on Environmental 2. B. C. Karkkainen, Columbia Law Rev. 102, 903 (2002). the statement produced. Quality, 722 Jackson Place, NW,Washington, DC 3. D. Bear, Nat. Resour. J. 43, 4 (2003). 3 4. J. F. DiMento, H. Ingram, Nat. Resour. J., in press. NEPA explicitly states that agencies are 20503, USA. Department of Geography, University of 5. N. N. Mccold, J.W. Saulsbury, Environ. Manage. 20 (no. 5), to use an interdisciplinary approach to their Alabama, Tuscaloosa, AL 35487–0322, USA. 4Newkirk 767 (1996). work, but this is not apparent in current EISs. The problem is that, despite years of effort and the development of university TECHNICAL COMMENT ABSTRACTS programs that claim to teach interdiscipli- COMMENT ON “Long-Lived Drosophila with Overexpressed dFOXO in Adult nary environmental research, the ability to Fat Body” perform it in the real world of deadlines and finite resources does not yet exist. Another Marc Tatar problem is the separation of “social” Giannakou et al. (Brevia, 16 July 2004, p. 361) reported that dFOXO overexpression extends Drosophila life-span. impacts from “environmental” impacts in However, mortality was similar among experimental and control groups in replicate trials, except for one week EISs. The underlying principle of NEPA is when there were many deaths in control adults.This early event reduces the control survivorship and is mistaken that all impacts of a project are eventually for evidence that aging is retarded by overexpression of dFOXO. Full text at www.sciencemag.org/cgi/content/full/307/5710/675a social, as they ultimately affect people. Evaluation of the cumulative effects of sev- RESPONSE TO COMMENT ON “Long-Lived Drosophila with Overexpressed eral projects is also missing from most EISs, which devote the majority of their dFOXO in Adult Fat Body” analysis to the current project (5). Maria E. Giannakou, Martin Gross, Martin A. Junger, Ernst Hafen, Sally J. Leevers, Linda Partridge Over the past 35 years, NEPA has Tatar suggests that the conclusions of our study are erroneous due to excessive mortality in young control flies.We greatly improved the quality of U.S. gov- show that,both in our published results and in other results we had at the time of publication,there is a highly repro- ernmental decisions regarding the environ- ducible increase in overall survival and maximum life-span of females with dFOXO overexpression in fat body. ment and enhanced public participation in Full text at www.sciencemag.org/cgi/content/full/307/5710/675b

PepChip® Peptide 1,152 protein Microarrays Custom Peptide kinase Synthesis

Protein substrates on Interaction Mapping PepChip Kinase is a novel peptide microarray designed for the analysis of protein kinase a chip! Epitope activity, ideal for assay development, Mapping drug discovery and signal transduction research.

Features and benefits Applications Peptide • Fast and complete insight in • Substrate profiling of known Lead Finding kinase substrate specificity and unknown kinases • Minimal use of valuable kinase • Specificity testing of kinase proteins inhibitors • Compatible with purified kinases • Cellular kinase activity profiling: and cell lysates kinomics

For further information, please contact: Pepscan 8219 PH Lelystad T +31 320 23 72 00 E [email protected] Edelhertweg 15 The Netherlands F +31 320 23 81 20 I http://www.pepscan.com

www.sciencemag.org SCIENCE VOL 307 4 FEBRUARY 2005 675 OOKS et al. B way back to the earliest recognizable forms of EVOLUTION life. Even for one accustomed to thinking about phylogeny and deep time, it is remark- Voyage to the Bottom of the Tree able to ponder that unbroken line of descent, stretching over billions of years and extend- Charles Francis Delwiche ing back to, well, primordial ooze. Men dreme alday of owles and of apes, By its very nature, an accumulat- And of many a maze therwithal. ing phylogeny of life on Earth lacks –Geoffrey Chaucer, The Canterbury Tales the focused clarity of Dawkins’s “self- ish gene”—one of the strongest iology is a science with many stories. metaphors in popular science and the Each organism is a unique product of subject of one of the most important Bits evolutionary history, its adaptation popular-science books ever (1). The to its environmental niche, and the con- Ancestor’s Tale is loosely modeled straints imposed by the laws of physics. With after Chaucer’s The Canterbury Tales, roughly 1.7 million known species, each one in which a group of pilgrims tell tales of which is enormously complex in its own on their journey to Canterbury. right, there is a bewildering array of curiosi- Dawkins uses this as an extended ties, exceptions, and oddballs. Given this riot metaphor, envisioning each branch of of biological diversity, it can hardly be sur- the tree of life as a group of pilgrims prising that there has been a trend toward the that joins us on our journey into the “model systems” approach, which picks a past. As each group joins the main handful of species to study in great detail. assemblage, after describing the After all, it would be a great accomplishment group and the reconstructed conces- to fully understand just one species. But tor, he relates stories that are inspired model systems can only take us so far; to by that group, using them to illustrate understand life it is necessary to grapple with fundamental biological principles. its full diversity, including that which is unfa- These tales are generally interesting miliar or unseen. In a very real sense, each and sometimes brilliant. With a couple species has its own tale to tell. of harmless exceptions, he wisely In The Ancestor’s Tale, Leading to lungfish and us. Dawkins identifies the resists the urge to tell these stories approaches the topic of biological diversity by common ancestor of lungfish and terrestrial vertebrates from the animal’s perspective, com- descending the tree of life one branch at a as a lobe-finned fish, the concestor reconstructed here. menting, “[others] might get away time, starting from the extant twig of six bil- with it, but not me.” The metaphor of a lion individual humans. At some point in the tor. Figures depict the topology of the growing pilgrimage really does help envi- not-too-distant past (perhaps 6 million years branches joining our own lineage, provide the sion relationships, and it throws into stark ago), our ancestors probable dates, and include tiny images of relief the difference between “relict” groups, The Ancestor’s Tale along with those of representative members of the descendant like monotremes (egg-laying mammals), A Pilgrimage to chimpanzees and . In the Weidenfeld and Nicolson edi- that consist of a few survivors of an ancient bonobos were mem- tion (which I read), these images are generally the Dawn of Life lineage and major groups, like sauropsids bers of a single beautiful photographs; in the Houghton (birds and living “”), that have under- by Richard Dawkins species, neither Mifflin version, they have been replaced with gone vast diversification. Weidenfeld and Nicolson, human nor chim- very simple line drawings. Readers of the U.S. The Chaucerian theme is cute and a lot London, 2004. 528 pp. panzee. Dawkins edition will also find, unfortunately, that it of fun, but it leads to titles of the individual £25. ISBN 0-297-82503-8. calls this species a lacks the often striking, full-page, color recon- stories such as “Prologue to the Galapagos concestor (for “com- structions Malcolm Godwin prepared of most Finch’s Tale” (an outstanding essay on the The Ancestor’s Tale mon ancestor”). He concestors—and that all of the illustrations it potential rapidity of adaptative evolution) A Pilgrimage to the traces our lineage does contain are presented in black and white. and “The Brine Shrimp’s Tale” (a highly Dawn of Evolution back through pro- The subject is timely. Although even a speculative essay on the origin of a dorsal Houghton Mifflin, Boston, gressively more decade ago the structure of the tree of life was versus ventral nerve cord). The tales’ titles, 2004. 687 pp. $28. ISBN 0- inclusive and stout largely a matter of speculation, huge 618-00583-8. unfortunately, often convey next to nothing branches to the three advances have been made in recent years, and about their content, a circumstance that - main trunks and the tree is now beginning to come into sharp mately proves rather confusing. finally to the root of the tree of life. Describing focus. Only a handful of the branch points Although in its entirety Dawkins’s narra- (and numbering) concestors, Dawkins identi- covered in the book remain in serious ques- tive follows an elegant arc, each individual fies 39 waypoints on this backward journey tion, and these are clearly identified. tale follows its own distinct path. Herein lies through time. At each of these “rendezvous,” Dawkins’s task was simplified by the fact that the book’s greatest problem, as many of the he stops to discuss the ancient common ances- humans are members of a relatively depau- tales wander a fair bit along the way. What is perate lineage that has been subject to inten- more, Dawkins distracts the reader with sive study. Other parts of the tree remain many marginal notes (often more than one The reviewer is in the Department of Cell Biology much more poorly understood, but for the and Molecular Genetics, University of Maryland, per page), some of which are quite extensive. College Park, MD 20742–5815, USA. E-mail: first time at least the fiber of our own ances- The effect is the textual equivalent of trying to

[email protected] try can be traced with some certainty all the keep track of the epicycles in a geocentric CREDIT: DESIGN MALCOLM GODWIN © MOONRUNNER

676 4 FEBRUARY 2005 VOL 307 SCIENCE www.sciencemag.org B OOKS ET AL. model of the solar system. Add to that the EVOLUTION Unfortunately, nearly all refer to the benefits cryptic titles of the tales and one becomes of knowing the phylogeny for a particular downright dizzy. Dawkins has written Seeing the Forest group and say little about those benefits that another wonderful book, but the manuscript can only come from assembling the entire would have benefited from a good, firm edit. for the Trees Tree. In addition to revealing common pat- Had the text been reduced by a third and the terns or coordinated evolution among clades, marginal notes either incorporated in the Scott J. Steppan having the whole Tree should lead to more main text or eliminated, The Ancestor’s Tale important but as yet unanticipated insights. could have become a true classic. It should ssembling the Tree of Life presents a For example, would Wegener have imagined also be noted that Dawkins is an entertaining preliminary view of one of the grand continental drift if he had only a collection of author precisely because he is not afraid to Aenterprises of modern science, resolv- road maps and no global map to work with? express opinions. However one might feel ing the phylogeny of all life. Imagine a vast We biologists need our own globe. about the particulars, at times these opinions tree whose myriad branches lead to millions Following the introductory section, 26 become downright caustic, and they trivialize of leaves. Each leaf, itself composed of innu- chapters by authorities on major branches the tales in which they appear. Conservative merable parts, represents an individual (clades) summarize the state of our phylo- readers might risk an aneurysm. species in the history of genetic knowledge. These begin at the base The taxonomic scope of The Assembling the life, and the tree stands of the Tree, where contributors highlight, Ancestor’s Tale is strongly affected by the Tree of Life billions of years tall. for example, the recent recognition that the author’s understandable decision to fol- Joel Cracraft and Revealing that tree is earliest branchings split life into three low human ancestry. We are animals, so Michael J. Donoghue, the shared vision of the domains: the bacteria, archaea, and eukary- this is fundamentally a book about ani- Eds. world’s systematists, otes. The chapters then proceed up the Tree mals (i.e., metazoans). In the one chapter but for now it remains a through smaller branches and less inclusive Oxford University Press, dream. We do not know on plants, Dawkins comments, “I ended a New York, 2004. 592 pp. groups (e.g., green plants, animals, and previous tale by remarking what delight it $59.95, £36.50. ISBN 0- what the whole “Tree of arthropods) to consider such “crown” is to be a zoologist at such a time. I could 19-517234-5. Life” looks like. We can groups as flowering plants, annelid worms, have said the same about being a botanist. only see parts of it, and and birds. In each chapter, the authors sum- What a pleasure it would be to demon- our situation is worse marize the constituent subgroups and typi- strate Deep Green [(2)] to Joseph Hooker— than that of the proverbial three blind men cally describe supporting evidence, regions in the company of his close friend Charles trying to describe an elephant. Thousands of of uncertain relationship, and definitive Darwin. I almost weep to think about it.” us work on particular branches, which are morphological features. Afterward, Don- Nonetheless, in practice Dawkins seems to hidden from one another in a mist. This vol- oghue and three other leading evolutionary follow another great philosopher of science, ume, the product of a 2002 symposium by biologists (Edward Wilson, David Wake, Tom Weller, who said, “The evolution of the same name held at the American and David Hillis) offer short summary per- plants is an important chapter in the history of Museum of Natural History (AMNH) in spectives. In the final chapter, the editors tie life. However, it’s a pretty dull chapter, so New York, seeks to blow away the mist and everything together by assembling a 138- we’ll skip it.” (3) Furthermore, to my - reveal the structure of the whole Tree and, in taxon synoptic tree. ishment, the Archaea—a third of all life—are doing so, galvanize the systematics commu- Taken individually, the chapters are use- not allotted a single tale. nity toward unifying its goals. ful summaries of our current understand- But so be it. Dawkins is an enormously A complete Tree of Life (hereafter ing, but they seem like disconnected limbs. talented author, and The Ancestor’s Tale is “Tree”) holds enormous promise for many Nonetheless, the Tree will start to assemble expansive, current, and authoritative. There fields of science, but the task of revealing it itself—an emergent property of the discon- are, of course, technical errors and dubious is an enormous undertaking—one that nected parts—in the minds of those readers assertions to be found. Few texts of such requires more data than the who take the time to read far enough. In that scope are without them. These flaws, how- Project (just one leaf on the Tree) and orders indirect way, the editors have met their goal. ever, are mostly minor, and the book avoids of magnitude more computation. Even small In addition, even the most broadly trained many pitfalls that have trapped other parts are difficult; as Michael Whiting notes, comparative biologists will discover unap- authors. It would be an excellent choice for “A child can tell a beetle from a wasp an undergraduate honors seminar in zool- from a butterfly, but even the entomo- ogy and could serve a graduate student well logically erudite is left pondering in preparation for oral exams. It is also which two insects are most closely entertaining, witty, and—at least in the related.” The volume, edited by lead- “Pilgrimage to the Dawn of Life” version— ing systematists Joel Cracraft beautifully illustrated. If it still leaves room (AMNH) and Michael Donoghue for the botanists and microbiologists of the (), begins with three world to present their perspective on the tree chapters that explain why assembling of life, who am I to complain? the Tree is important to science and society. Most of the reasons offered References and Notes will be familiar to biologists, as the 1. R. Dawkins, The Selfish Gene (Oxford Univ. Press, revolution in systematics has pene- Oxford, 1976). 2. “Deep Green” refers to a project to reconstruct the trated many different fields. phylogeny of green plants and to a hyperbolic visuali- zation of that phylogenetic tree, available at http://ucjeps.berkeley.edu/map2.html. The reviewer is in the Department of 3. T. Weller, Science Made Stupid (Houghton Mifflin, Biological Science, Florida State University, Boston, 1985). Tallahassee, FL 32306–1100, USA. E-mail:

CREDIT: DAVID SACKS/TAXI/GETTY IMAGES CREDIT: SACKS/TAXI/GETTY DAVID 10.1126/science.1105582 [email protected] Iconic metaphor.

www.sciencemag.org SCIENCE VOL 307 4 FEBRUARY 2005 677 B OOKS ET AL. treatment of the debate over the BROWSINGS effects of long-branch attraction on The Elements. Earth, Air, Fire, and analyses of the relationship Water. Art Wolfe,text by Craig Childs. between the fly orders Strepsiptera Sasquatch, Seattle, WA, 2004. 176 and Diptera (2). A more pervasive pp. $45. ISBN 1-57061-405-9. and subtle, yet profound, conse- In ancient Greece, philosophers quence of this methodological bias held that the physical world around is omission from most chapters of them was derived from four ele- fundamental aspects of evolution- ments: earth, air, fire, and water. ary history, like timing of events Wolfe presents a collection of his and rates of diversification. The color landscape and nature photo- emphasis in the volume is entirely graphs that explores the diversity of on the sequence of branching. forms these substances take. There (Branch lengths are not important are sheer rock walls of the Karakoram in parsimony analysis, and their Range, Pakistan, and loose sediments estimates are generally unreliable. of the Colorado River’s subaerial In contrast, they are integral to delta plain, Baja California (left); model-based methods.) As a result, morning mists and midday clouds; the tempo and mode of evolution volcanic eruptions and flaming (3) are lost, and we cannot see forests; Hawaiian breakers and whether the Tree looks like a Antarctic ice. The four elements are spreading oak, a willow, or a bam- also used as background for portraits boo grove—we have little sense of of wildlife, flowers, and trees. Many its gestalt. The lack of resolution in of the images are carefully composed some parts of the Tree is therefore to capture patterns of light and con- attributed to a lack of data rather trast. In four short essays, Childs than to the much more interesting offers his impressions of the effects possibility of rapid diversification. the elements have on humans and Branch lengths—as indicators of the natural world. time or amount of evolution—are important to almost every aspect of comparative biology, and the vol- preciated diversity in less familiar groups its revelation of the patterns among diverse ume would have benefited from the more and the kind of fascinating organisms that clades. Many authors cite the explosion of nuanced vision their consideration would inspired many of us to become biologists. molecular data as the reason for the revolu- have offered. These benefits would have been even easier tion in phylogenetics, especially for the The summary chapters praise the to appreciate if the material was presented field’s transformation since the previous progress and promise more to come. I in the more dynamic and immersive experi- symposium that attempted to view phyloge- would have preferred a more critical analy- ence of the volume’s Web analog, the Tree netics across all of life (1), held in 1988. The sis of the overall state of this resource- of Life project (http://tolweb.org/tree/). (It most publicized cases of conflict between limited field. Where are the biggest holes? is a shame that updatable, peer-reviewed molecules and morphology are not repre- Should we focus on broad taxonomic cover- Web pages still lack the professional status sentative of that revolution: The tidal surge age of a few universal genes, overlapping of static book chapters.) of molecular data seems to have confirmed sets of many genes, or perhaps new initia- Most authors have taken their charge numerous old hypotheses while rejecting a tives to train morphologists? But in the end, very seriously and have written unbiased, few but, most importantly, resolving many the big picture emerges from the details, synthetic, and useful accounts. Particularly branches that morphological evidence did and we gain a better appreciation of how the readable chapters include those on not. One is struck by the great reliance on a branches fit together and where some of the Holometabola (insects characterized by single gene—the small subunit (SSU) of the bigger questions remain. The vision complete metamorphosis), land plants, and ribosomal DNA, also known as 18S—for Cracraft and Donoghue articulate in their (vertebrates, hagfish, lancelets, most resolution deep in the Tree, even introduction does emerge from the mist, and tunicates). A minority of the contribu- within phyla. Elsewhere, despite frequent incomplete though it may appear. tors have yielded to provincialism, focusing accolades to molecular data, the recognition Assembling the Tree of Life should also on their own work or dismissing informa- of many clades (especially among chor- meet the editors’ larger goal. It will help the tion (e.g., molecular) that they distrust. The dates) continues to rely on morphology. systematic community aspire toward a most extreme position appears in the mam- The other broad impression the volume common goal, identify priorities for future mal chapter, whose authors eschew the leaves is that of an imbalance toward coordinated work, and mobilize our summary format in favor of lecturing on authors who favor parsimony for phyloge- resources. their preferred systematic procedures. Only netic analysis over model-based or sta- a handful of conflicting conclusions appear; tistical methods such as likelihood. References and Notes

one is the description of the Holometabola Individually, this imbalance is not very 1. B. Fernholm, K. Bremer, H. Jörnvall, Eds., The Hierarchy THE ELEMENTS of Life: Molecules and Morphology in Phylogenetic as a group whose monophyly is either rou- important because all chapters include Analysis (Nobel Symposium 70, Elsevier, Amsterdam, tinely supported by both morphology and authoritative authors. The reviews of find- 1989). 2. J. P. Huelsenbeck, Syst. Biol. 46, 69 (1997). molecules (Whiting) or never supported in ings by other researchers are generally fair, 3. G. G. Simpson, Tempo and Mode in Evolution any molecular data (Rainer Willmann). although occasionally conflicting model- (Columbia Univ. Press, New York, 1944).

The volume’s principal utility stems from based results are brushed aside—as in the 10.1126/science.1106586 CREDIT:ART WOLFE FROM

678 4 FEBRUARY 2005 VOL 307 SCIENCE www.sciencemag.org ESSAY GLOBAL VOICES OF SCIENCE It Takes a Village: Medical Research and Ethics in Mali Ogobara K. Doumbo

The world is populated by rich and poor tion, and showed compassion for their suf- of ethics contin- nations filled with a diversity of peoples and fering. The patient and the caregiver in this ued when I started my customs. For those scientists and physicians care-providing system are so intimately con- own medical research career in 1984. I was trying to have an impact on global health, nected that my grandfather considered the fortunate at that time to have been at the conducting clinical trials and other types of patients as part of his family, sharing the University of Marseille where I worked with medical research in these varied places, and same food and shelter. Because of my grand- parasitologist Philippe Ranque in the first among these different cultures, is central to father’s fame, and because he specialized in Phase II trials in Mali of ivermectin, then a the cause. It also is imperative two diseases—pharyngitis and new drug for the treatment of onchocercia- that these efforts are undertaken breast tumor/infection—some sis, or river blindness. Like my grandfather, in ethical ways that respect and This yearlong patients came to him from very Dr. Ranque’s priority was to show his honor those many individuals essay series remote areas. patients attention, care, and compassion. Dr. and communities that agree to celebrates 125 The respect my grandfather Ranque believed that protection of volun- participate in the investigations. years of Science by showed while interacting with teers in medical trials was more important To do that, researchers need to inviting researchers his patients marked my life. At than achieving scientific goals. develop an understanding of the from around the the age of 10, I decided to Over time, my exposure to this sort of unique ways in which different world to provide become a medical doctor, and 7 ethics-in-the field, as well as to more aca- cultural groups make decisions. a regional view of years later I began my medical demic discussions of ethics, convinced me Only then can the investigators the scientific studies at the National School of that scientists must consider ethical issues feel assured that the human enterprise. Medicine and Pharmacy in throughout the entire research process, from beings who consent to partake in Series editor, Bamako, Mali. There, in my sec- identifying a research question to analyzing their studies are fully informed Ivan Amato ond year of medical studies, dur- and interpreting data. about the possible risks and bene- ing rounds with the late Bernard In Mali, where most of the population fits of such participation. Duflo of the Internal Medicine cannot read and where the social structures Without knowing it at the time, I began to ward of the National hospital “Point G,” I demand that decision-making be done more develop a sensibility to these issues as an 8- witnessed a similar intensely ethical attitude communally than in the West, we have had year-old child when I used to follow my in the patient-physician relationship. My to develop new procedures for obtaining grandfather while he practiced traditional grandfather and Dr. Duflo, a French physi- informed consent from participants in our medicine in the Dogon country in northeast cian, were from two completely different malaria research. The process doesn’t al- Mali. I was impressed by his attitude toward cultures, but they shared the same attitude in ways work, takes a long time, and requires his patients. He was very close to them, healing patients. more discussion and leg work than in the talked to them with respect and considera- My “indoctrination” in the importance West, but it generates mutual understand-

Ogobara K. Doumbo Mali Ogobara K. Doumbo, director of the Malaria Research and Training Center at the University of Bamako, knew he would become a doctor at the age of 10. Immersed initially in the traditional medicine of his grandfather, he earned his medical credential in his home country and then later earned a Ph.D. in tropi- cal medicine and parasitology at the Universities of Marseille and Montpellier in France.With expertise also in tropical medicine and parasitology, and with extensive research connections throughout the world, Dr. Doumbo has been making a global impact on the study, treatment, and prevention of malaria. With both in-the-field and academic exposure to medical ethics issues, he also has been helping to develop research protocols that respect and honor the diverse populations that become involved in his medical research. In 2003, following a nomination by the president of Mali, Dr. Doumbo became a mem- ber of his country’s National Committee on Ethics.He also has been knighted by France’s Legion of Honor and Mali’s National Order. His center was selected in 2003 as a Center of Excellence in Clinical Research by the Agence Universitaire de la Francophonie (AUF) and as a malaria vaccine site by the National Institute of Allergy and Infectious Diseases of the National Institutes of Health in Bethesda, Maryland.

All essays appearing in this series can be found online at www.sciencemag.org/sciext/globalvoices/ CREDIT: OGOBARA DOUMBO

www.sciencemag.org SCIENCE VOL 307 4 FEBRUARY 2005 679 G LOBAL V OICES OF S CIENCE

a dynamic approach erates inside the insects. Such direct exposure to obtaining informed to these mosquitoes—even ones that were consent and to main- themselves not yet carrying the parasite and taining it over time. grown under controlled conditions—generates Usually, this was many ethical concerns. How could we prove to a stepwise process. ourselves and to the community that the lab- First, we needed to reared Anopheles really were safe? How get permission from should we select the volunteers? How should the community to we approach the community and document the proceed. This began process of community permission and individ- as a discussion with ual consent (versus assent)? How should we the group of village explain the malaria life cycle to a population elders. Next, we con- where 70 to 80% of its members are illiterate? vened focused group The first time we presented our protocol to discussions with the the Institutional Review Board (IRB) of the heads of extended Faculty of Medicine, Pharmacy and Odonto- families. Then we stomatology, which oversees informed consent “Wabu” is a term that traditional healers in Mali use when refer- held similar discus- procedures, and to the village community, the ring to a condition marked by fever, seizures, delusions, or coma, sions with mothers design was rejected. The IRB deemed that it the hallmarks of severe malaria.Traditional thought assigned the whose children might was not acceptable to expose humans to cause of wabu to a bird taking the spirit of a child at the moment become part of the Anopheles even if the insects were reared in a that the bird and the child cry out simultaneously. malaria study. Finally, well-qualified laboratory. We had to spend we obtained consent days and nights explaining the protocol to the of the individual fam- villagers and to the Malian IRB members. Part ing, medical care for the participants and ilies involved in our cohort. The consent of the process involved bringing representa- often their entire communities, and valuable process was open and better suited to the tives from two villages to visit our center at scientific results.* needs of the population than were more con- Bamako so that they could see how the expo- ventional approaches. It generated more con- sure procedure would unfold. They spent 2 Ethics in the Field: Informed Consent fidence by the villagers in the research project days with us and questioned us about all At the Malaria Research and Training Center and a better understanding for us of the village aspects of the experiment. In the end, the pro- (MRTC) of the University of Bamako, Mali, culture and behavior. In developing this tocol was accepted, providing testimony to the we conduct different types of research, includ- approach, I always had my grandfather and ways in which responsible, open, and patient ing laboratory-based studies, field research, other ethics mentors in mind. explanation of well-designed studies some- and clinical trials. For us, ethical considera- The villagers in the study received a tangi- times can alleviate participants’ fears and tions constitute a dynamic process and are an ble benefit too. Because we needed to exam- misgivings.† integral part of the research enterprise. Here, I ine the children many times during the course The third NIAID-funded project—in col- will focus on some study designs used at the of the study, it meant that the villagers bene- laboration with colleagues at the U.S. Centers MRTC and how we incorporate ethical con- fited from repeated contacts with the study for Disease Control in Atlanta, GA, and the siderations in our procedures for obtaining physicians who lived in the village. Previous National Malaria Control Program of Mali— informed consent. I will also compare the malaria studies in Africa commonly used a was designed as a supplement to the original strategies for getting documented and signed design typical of developing TMRC project. It was informed consent in developing countries, on countries in which the interaction particularly ethically the one hand, and in North America or Europe, between study subjects and inves- charged because it in- on the other. tigators is more limited and usu- volved a vulnerable In recent years, we conducted an observa- ally confined to clinical settings. population of preg- tional study of the natural history of malaria The second TMRC project was nant women. We in the field as part of the Mali-Tulane Tropical also unique in the strategy of get- wanted to conduct this Medicine Research Center (TMRC) project, ting informed consent. We wanted study to generate data funded by the National Institute of Allergy to test gametocyte infectivity in important for develop- and Infectious Diseases (NIAID). The grant the community by directly expos- ing poli- included three projects all located at ing members already carrying the cies to help protect Bancoumana, a large rural village in Mali gametocytes to Anopheles gam- pregnant women from with 12,000 inhabitants. biae mosquitoes—the most effec- contracting malaria in In Project 1 we studied a cohort of about tive vector of the malaria parasite, ways that minimize 3500 children under 9 years of age, all of Plasmodium falciparum, in risks to the fetus.‡ whom had been exposed to malaria. We Mali—collected in that village. We designed a recorded data on the natural history of Our aim was to determine the fac- community-based malaria with the aim of identifying risk fac- tors that influence how efficiently open label trial to tors for infection or disease. the mosquitoes would pick up the compare three pro- Because our studies were being done in the gametocytes from the infected vil- phylactic drug regi- field and in a variety of social structures that lagers to initiate the sporogonic Champion Infector: The Anopheles mens in pregnant respond differently to medical researchers cycle during which the infective gambiae mosquito is the most women. Given the

coming into their lives, we needed to develop form of the malaria parasite prolif- effective vector of malaria in Mali. special importance of A SOUDERS/CORBIS;WORLD ORGANIZATION HEALTH PAUL CREDITS: BOTTOM) TO (TOP

680 4 FEBRUARY 2005 VOL 307 SCIENCE www.sciencemag.org G LOBAL V OICES OF S CIENCE

pregnant women in African culture, we knew believed that “they gave their words” and that children and pregnant women. It can be espe- that we needed to be extremely careful about that should be sufficient. It took very careful cially challenging to convince the community ethical considerations. Although these drugs explanation and patience to overcome this that it ultimately is in its best interest to allow were recommended by the National Malaria resistance. Informed consent must be based children to participate in trials that could have Control Program and the World Health on a thorough understanding of the society in widespread medical benefits in the future. Organization, their use in pregnant women which the study is to take place. For outsiders, One way to earn the confidence of the needed rigorous ethical scrutiny. the role of local guides, local investigators, community is to provide medical care for the Central to the project, of course, was to and socio-anthropologists is critical. community while the study is being con- obtain the community’s permission to conduct Documents and legal language also pose ducted. In rural regions of developing coun- the study. A key early step was to discuss the difficulties. One of them is that in developed tries where medical care is limited or nonex- benefit of learning more about these drugs with countries, the heavy use of legal language istent, the research team often has to set up the women’s local council. Then we met with and documents makes it increasingly diffi- its own clinic. Providing standard care for the mothers-in-law of each pregnant woman. cult and murky for participants to discern the both study participants and others in the After all of these discussions, we met with the risks and benefits of participating in studies. community during a research project where individual pregnant woman. Toward the end of The goal of these legal documents seems to the team is the sole source of medical care the process, the women requested that we meet have more to do with protecting the investi- can be a form of community compensation. with their husbands and fathers-in- Almost all studies in law as well. (It is a widespread cus- developing countries tom in African cultures that a preg- should guarantee care nant woman belongs to the family- for volunteers who in-law, which is responsible for her experience serious ad- care.) The overall goal of this proj- verse events during ect was to determine risk factors the study and after for contracting malaria, and we it has been completed. were able to offer the community Care must be taken to the benefit of yearly feedback of ensure that the provi- all the results about the likely sion of these services, infectivity of their local mosquito and the establishment population, before the next trans- of a clinic, do not mission season. induce the commu- nity to participate in Consenting Partners the study when in fact These three experiences show it might not be in the how different the informed con- community’s interest sent process needs to be in differ- to do so. This ethical ent places in the world. For one dilemma is not yet thing, the issue of who “signs Unconsenting hosts. Women of child-bearing age, like these three walking to market in solved and remains off ” on consent must be carefully the Dogon region of Mali, are particularly vulnerable to malaria. a big concern. considered. Informed consent in The challenge of Europe and in North America usually gators and sponsors than the volunteers. obtaining informed consent for medical involves written documents, which the What’s more, legal language is hard enough research from communities in developing prospective volunteer must read and sign in for a highly educated person to understand. countries can be daunting, especially from order to participate in a study. The emphasis In Mali, less than 20% of adults are literate, rural and mostly illiterate populations in rural is on the autonomy of individuals. so written documents can easily discourage regions. Even so, our 15 years of international In some developing countries, however, rural populations from participating in stud- research experience in dealing with these individual and community consent are part of ies. Also, for rural communities, paper often complex issues of informed consent in Mali the same process. We cannot separate them in means trouble with the government. shows that these difficulties can be overcome our countries, and this reality should be From both ethical and biological stand- in a way that benefits medical science and understood by sponsors and funding agencies points, the particular case of pregnant women public health. and northern research institutions. We need to needs special attention. In Malian traditional think about the protection, safety, risks, and communities there is a very good but discreet Notes *O. Doumbo, Multilateral Initiative on Malaria (MIM), benefits of individuals and of the community representation of women in decisions con- Durban, South Africa, March 1999. at large. We have learned in particular that the cerning the village’s affairs. Foreign re- †Y.T.Touré et al., Am. J. Trop. Med. Hyg. 59, 481 (1998). initial focus and discussion should be with the searchers may overlook this important ‡K. Kayentao et al., J. Infect. Dis. 191, 109 (2005). leaders of the community, rather than individ- involvement of women and thereby lose the I thank all the populations that participated in the studies, uals. By approaching individuals first, one is confidence of the community. Similar deci- the MRTC/DEAP teams and partners, and my collaborators likely to introduce social conflict in the vil- sion-making subtleties go with enrolling who read and corrected the manuscript: M. A. Thera, A. A. Djimdé, and D.A. Diallo. lage, and this could be unethical. children in medical trials. Another challenge is the need to document In the developed countries, children nor- The author is in the Department of Epidemiology of the consent process using a signed document. mally are excluded from medical trials of new Parasitic Diseases (DEAP), Faculty of Medicine, Pharmacy At the beginning of our TMRC projects, the medicines, but in Africa we have to test prod- and Odonto-Stomatology, University of Bamako, Mali, BP villagers we approached were opposed to ucts on children because the target populations 1805, Bamako, Mali. E-mail: [email protected]

CREDIT: VANNINI/CORBIS SANDRO signing any document, because they strongly for malaria and many infectious diseases are 10.1126/science.1109773

www.sciencemag.org SCIENCE VOL 307 4 FEBRUARY 2005 681 PERSPECTIVES OCEAN SCIENCE approach allows, for example, the strengths of El Niño conditions to be assessed for times when ocean data were largely The Ocean’s Seismic Hum unavailable. Other researchers are combining remote Sharon Kedar and Frank H.Webb sensing data from ocean satellite altimeters and data from the global seismic network to he first seismometers capable of global ocean-wave observations and mod- investigate global interactions between the measuring ground vibrations with els has led to new insights into the origins of oceans and the solid Earth. Rhie and Tperiods of several seconds were ocean microseisms and new Earth science Romanowicz (7) have discovered that the installed in the early 20th century. Since applications of these signals. ocean’s hum extends to periods of several then, the devices have recorded a continuous Potential oceanographic applications minutes and have shown that background seismic hum, called “ocean microseisms.” were considered as early as the 1960s, when vibrations of the solid Earth are excited in This hum is not the Haubrich et al. showed that storms can be the northern Pacific ocean in Northern Enhanced online at result of tectonic located and tracked with seismic data (4). Hemisphere winter, and in southern oceans www.sciencemag.org/cgi/ forces, but rather This method has been improved upon in in the summer. Ocean microseisms thus act content/full/307/5710/682 the response of the recent years. Bromirski and Duennbier (6) as a meter by which global-scale ocean solid Earth to have shown that the wave climate (that is, activity can be monitored. ocean wave-wave interactions, which have the ocean conditions that give rise to and Microseisms are also used to probe an annual global cumulative seismic energy sustain ocean waves) can be reliably recon- Earth’s structure through seismic tomogra- comparable to that from earthquakes. structed with archived seismic data. This phy. Crustal-scale tomography typically Long considered noise by seismologists, ocean microseisms have recently been found to be a useful resource for the inter- disciplinary study of our planet. They pro- 60° vide a record of the state of the oceans since the early 20th century and are a passive seismic source for probing the geological 40° structure of Earth’s upper crust (0 to 20 km). Origins, applications, and future stud- 20° ies of ocean microseisms were discussed at the Fall 2004 meeting of the American Geophysical Union (1). 0° Early studies showed that ocean micro- 17 January 1998, 3 hrs GMT seism signals are linked to ocean swell condi- –20° tions (such as wave direction, amplitude, and 150° 200° 250° 300° 350° period) but have half the swell period. The debate over their origins was settled by Longuet-Higgins, who showed in 1950 that 60° the interaction of water waves with similar frequencies but opposing directions generates 40° a second-order pressure wave with half the period and an amplitude proportional to the 20° product of the wave heights (2). Unlike the Latitude Latitude pressure field generated by traveling waves, this pressure wave does not wane with depth 0° and efficiently couples with the solid Earth to 23 January 1998, 6 hrs GMT generate seismic surface waves. –20° The conditions for generating these 150° 200° 250° 300° 350° wave-wave interactions probably occur at Longitude cyclonic depressions and along steep coast- lines. Until recently, information about the 012345678910 source of ocean microseisms was inferred Significant wave height (m) from laboratory experiments, analysis of seismic data, and sporadic ocean observa- Ocean microseisms direction at Lodore seismic array, Wyoming tions from buoys and ocean-bottom pres- Ocean microseisms direction at Anza seismic array, California sure sensors (3–5). In recent years, the com- Tracking wave-wave interactions. The maps show global wave heights [from the NOAA Wave bination of seismic data and improved Watch III model (12), see color scale at bottom] and arrival directions of ocean microseisms at U.S. seismic arrays (from seismic data; colored arrows). (Top) Microseisms recorded in Wyoming are The authors are at the Jet Propulsion Laboratory, 4800 dominated by wave-wave interactions near the British Columbia coast, and those recorded in south- Oak Grove Drive, Pasadena, CA 91109, USA. E-mail: ern California by interactions off the coast of Baja California. (Bottom) A North Atlantic storm swell

[email protected], [email protected] hitting the steep Labrador coast triggers transcontinental microseisms. CREDIT: BOULDER V.AT OF COLORADO SCHULTE-PELKUM/UNIVERSITY

682 4 FEBRUARY 2005 VOL 307 SCIENCE www.sciencemag.org P ERSPECTIVES employs energetic, sporadic sources such as locally in coastal regions. However, several based measurements, is promising. This large explosions or earthquakes. Because times a year, cross-continental seismic joint effort is likely to benefit ocean and ocean microseisms are almost continuous, waves are excited when swell is reflected solid Earth science alike. they offer the possibility to monitor tempo- off a coast that is prone to ocean microseism ral changes in the crust (induced, for exam- generation (10). Rodriguez et al. (11) have References ple, by tectonic stresses and fluid flow) on proposed that microseism-generating 1. Ocean Microseisms: Observations and Applications, American Geophysical Union Fall 2004 meeting, 13 to much shorter time scales than is possible regions both in the open ocean and in 17 December 2004, San Francisco. with sporadic sources. Suggested in 1964 coastal regions should be directly observ- 2. M. S. Longuet-Higgins, Philos. Trans. R. Soc. London Ser. A 243, 1 (1950). (8), this method has become a reality with able and identifiable from space- and air- 3. R. I. B. Cooper, M. S. Longuet-Higgins, Proc. R. Soc. the proliferation of digital seismic data. At borne instruments. They pointed to syn- London Ser. A 206, 424 (1955). the meeting, Stehly et al. (9) showed that thetic aperture radar as the prime candidate 4. R. A. Haubrich, W. H. Munk, F. E. Snodgrass, Bull. Seismol. Soc. Am. 53, 27 (1963). ocean microseisms can be used reliably for for identifying regions of interaction 5. K. Hasselmann, Rev. Geophys. 1, 177 (1963). tomographic imaging. between swells of similar periods and 6. P. D. Bromirski, F. K. Duennebier, J. Geophys. Res. 107, 2166 (2002). To better understand and use this opposing directions, which are necessary 7. J. Rhie, B. Romanowicz, Nature 431, 552 (2004). ocean–solid Earth interaction, better for the excitation of ocean microseisms. 8. M. N.Toksoz, Geophysics 29, 154 (1964). knowledge of the state of the ocean wave Ocean wave-wave interactions and the 9. L. Stehly, N. M. Shapiro, M. Campillo, M. H. Ritzwoller, Fall AGU 2004, abstract S13C-1072. field that generates the ocean microseisms seismic energy they generate have largely 10. V. Schulte-Pelkum, P. S. Earle, F. L. Vernon, Geochem. is required. Combining seismic observa- been forgotten by oceanographers and Geophys. Geosyst. 5, 10.1029/2003GC000520 (2004). tions and wave action models, Schulte- swept aside as noise by seismologists. The 11. E. Rodriguez, S. Kedar, Fall AGU 2004, abstract S11C-04. Pelkum et al. (10) have analyzed oceanic emerging interdisciplinary effort of the two 12. H. L.Tolman, User manual and system documentation source regions and conditions for ocean communities to understand and use this of WAVEWATCH III, version 1.18, Tech. Rep. Ocean Modeling Branch Contribution 166, MOAA/NWS/ microseism generation (see the figure). fundamental interaction, taking advantage NCEP, 1999. Typically, ocean microseisms are generated of the riches of modern ground- and space- 10.1126/science.1108380

GENETICS equilibrium, real associations would be expected only between close markers and the A Century of Corn Selection trait of interest. An analysis of variance that fitted each William G. Hill marker locus individually and in pairs in sep- arate analyses of inbred and hybrid data sing conventional selection methods, Divergent selection in separate lines for showed that both dominance within loci and plant and animal breeders have made kernel protein concentration gave similar epistatic interactions between pairs of loci Umany beneficial changes to the yields responses, except that the low line reached were weak relative to additive effects. This and composition of crops and livestock (1). a plateau at about 5% protein (3). means that the oil concentration in heterozy- Yet we know little about the numbers, effects, To explain the large response in terms of gotes was intermediate between that of the and mode of action of the genes that account changes at the level of individual gene loci, two homozygotes, and that effects at different for these long-term changes. A recent paper Laurie, , and colleagues from the gene loci did not interact. The high correla- about maize selection in the journal Genetics Monsanto Company and the University of tion (0.75) of QTL effects on oil content esti- demonstrates that such information is slowly Illinois recently reported an analysis of the mated from inbred and hybrid plants is a fur- becoming available (2). maize data that itself took much time and ther indication that they act additively. With Since 1896, in one of the longest experi- work (2). Their goal was to identify quantita- the use of a stepwise multiple regression ments ever, biologists at the University of tive trait loci (QTLs)—regions of the genome analysis to select markers linked to QTLs and Illinois have continuously selected maize where genes influence the trait—by testing to account for linkage disequilibrium (corn) to change the oil composition of its the association between markers and the trait, between them, 50 markers were selected for kernels (1, 3). Separate maize lines have a standard technique of QTL analysis (4). the inbred data and 39 for the hybrids (where been selected for more than 100 genera- A cross between high and low oil-produc- differences are smaller). Significant effects tions according to whether the kernels con- ing maize lines from generation 70 was ran- were found on all 10 , with tain high or low amounts of oil, a trait of domly bred for 10 generations in a large pop- some clustering in the genome. agronomic importance (3). Typically, mean ulation (2) to reduce the effects of linkage dis- A major problem in QTL analyses com- oil concentration was estimated in 60 or so equilibrium. From each of 500 inbred lines prising many tests of significance is to com- ears (cobs) of maize, and seeds from only subsequently derived by self-pollination, promise between declaring false associations 12 were selected to propagate the next gen- DNA was extracted for genetic analysis and while missing real ones. To assess their find- eration. The change in oil concentration oil concentration was estimated in the ears of ings, Laurie et al. simulated data with effects was almost continuous (3) and substantial: inbred plants and of hybrid plants obtained by distributed similarly to what they observed, From a base of about 5%, the high oil-pro- outcrossing. The single-nucleotide polymor- and subjected these data to the same analysis. ducing line now has about 20% oil in the phisms (SNPs) chosen as markers of whether They concluded that they had detected about kernel, and the low oil-producing line has a region of the genome came from either the 63% of the QTLs and that about 33% of almost none (see fig. S1). The two maize high or low oil-content line differed substan- markers selected were not QTLs. Conse- lines differ by about 32 standard deviations tially in allele frequency between the lines or, quently, they calculated the correct number of (SD, 0.42% in the base population). exceptionally, filled gaps in the genetic map. QTLs to be about 50. After eliminating markers with strong linkage The estimated effects of the QTLs on oil The author is in the School of Biological Sciences, disequilibrium, the investigators focused their concentration were all much less than the line University of Edinburgh, Edinburgh, EH9 3JT, UK. E- analysis on 440 SNPs. Because markers more divergence of 15% at generation 70. Indeed, mail: [email protected] than 20 cM apart were essentially in linkage the largest had an effect (half homozygote

www.sciencemag.org SCIENCE VOL 307 4 FEBRUARY 2005 683 P ERSPECTIVES difference) of about 0.3% oil, and most had a responses to artificial selection. It is moot as geneticists to identify the genes and the difference of 0.1 to 0.2%. Only about 80% of to what defines a “small” effect, however. A molecular changes in them that cause these the QTLs had effects in the same direction as maize line containing 0.2% oil in the kernel many small but important differences in the selection response (38 of 50 inbreds, 33 of represents a difference of almost 1 SD quantitative traits. It is these small differ- 39 hybrids). For the others, the marker allele between homozygotes in the maize base pop- ences that generate variability in popula- from the high oil-producing line was associ- ulation; the largest effects detected in the tions, providing fuel for change through the ated with low oil—however, alleles of small chickens were almost as big (the variance in action of natural and artificial selection. effect can be fixed by chance in the opposite the F2 was much higher than in the base). The direction to that of selection if few parents are continuing responses to selection, therefore, References 1. J. Janick, Ed., Reviews, Volume 24, Part selected (5). The detected QTLs, if segregat- are not likely to be due mainly to continuing 1: Long-Term Selection: Maize. Part 2: Long-Term ing independently, could account for about tiny changes in gene frequency predicted by Selection: Crops, Animals, and Bacteria (Wiley, Hoboken, NJ (2003). half the genetic variance of the trait in the the infinitesimal model; instead they may be 2. C. C. Laurie et al., Genetics 168, 2141 (2004). population and, by summing their effects, due to the fixation of genes, including those 3. J. W. Dudley, R. J. Lambert, Plant Breed. Rev. 24 about half the divergence between the high arising by mutation after selection started (part 1), 79 (2004). 4. M. Lynch, B. Walsh, Introduction to Quantitative and low oil-producing maize lines. The (12, 13), which have appreciable effects Genetics (Sinauer, Sunderland, MA, 1998). remaining QTLs are likely to have similar or while segregating. The biological processes 5. A. Robertson, Proc. R. Soc. London Ser. B 153, 234 (1960). smaller effects on oil concentration because leading to oil concentration or chicken 6. L.Andersson, P. B. Siegel, personal communication. those with large effects were unlikely to be growth are obviously highly interactive, but 7. E. A. Dunnington, P. B. Siegel, Poultry Sci. 75, 1168 missed given the degree of genome coverage genes that contribute to selection response (1996) 8. A. Sewalem et al., Poultry Sci. 81, 1775 (2002). and the size of the experiment. must differ in effect when averaged over all 9. D. J. deKoening et al., Genet. Res. 83, 211 (2004). Similar findings have also been obtained other segregating genes. This may explain 10. S. Kerje et al., Anim. Genet. 34, 264 (2003). 11. B. Hayes, M. E. Goddard, Genet. Sel. Evol. 33, 209 in a recent study on divergent selection lines of the Laurie et al. finding that their detected (2001). poultry. Andersson and colleagues (6) ana- QTLs had approximately additive effects on 12. W. G. Hill, Genet. Res. 40, 255 (1982). lyzed a large intercross of poultry lines that oil production in maize. We have yet to dis- 13. B.Walsh, Plant Breed. Rev. 24 (part 1), 177 (2004). had been selected by Siegel in Virginia (7) for cover how such QTLs work, but several of 10.1126/science.1105459 high and low body weight (at 8 weeks of age) the SNPs associated with oil concentration Supporting Online Material www.sciencemag.org/cgi/content/full/307/5710/683/ for 40 generations and that differed in body were at candidate loci (2), so there are oppor- DC1 weight by a factor of about 9. Although 13 tunities to find out. It is a challenge for Fig. S1 QTLs were detected, none individually accounted for more than 3% of the body- ECOLOGY weight variance in the F2 generation, and each of these QTLs contributed only a small part of the divergence between the selected lines. Furthermore, the QTLs mainly had additive Untangling an Entangled Bank effects on body weight, as Laurie and col- David Storch, Pablo A. Marquet, Kevin J. Gaston leagues found in their maize analysis. In most other studies, however, QTLs of iodiversity, the most conspicuous prop- bushes, with various insects flitting about, substantial effect have been detected— for erty of life, has fascinated generations and with worms crawling through the damp example, QTLs for body size in poultry, not Bof ecologists and evolutionary biolo- earth, and to reflect that these elaborately only in broiler × layer (8) crosses, but also in gists. Part of this fascination arises from the constructed forms, so different from each commercial broiler populations still segre- fact that only a small fragment of this diver- other, and dependent on each other in so gating under intense selection (9). Some sity has been described and catalogued, pro- complex a manner, have all been produced by QTLs exerting large effects on the trait of viding endless opportunities to speculate laws acting around us” (1). interest found in mapping experiments have about the rest. Part arises from the sense that Some general patterns in contemporary subsequently been identified as a single any regularities and general patterns in the biodiversity do exist (2), and some of them are causative mutation—such is the case with biodiversity that we see today exist despite, or surprisingly simple. Take the species-area the mutation in the gene encoding insulin perhaps because of, the complexity of the relationship, for example. The number of growth factor–2 (IGF-2) in the pig, which processes and interactions that have driven species increases with the area sampled, often alters muscle growth in these animals (10). the dynamics of biodiversity through time linearly on a log-log scale, suggesting scale Although such effects are real, effects of and space. In the Origin of Species, Charles invariance. Such invariance is a prime area of QTLs declared significant tend to be biased Darwin made a specific appeal to this idea investigation among those interested in com- upward, and those of small effect are more when he wrote his famous description of the plex systems. Many phenomena—ranging likely to be missed (4). Models of the under- complex ecology of a bank covered by dense from the frequency distribution of species lying distribution of gene effects indicate an vegetation: “It is interesting to contemplate extinctions to allometric relationships exponential form, with numbers increasing an entangled bank, clothed with many plants between body size and rates of various bio- as effects get smaller (11). Too much varia- of many kinds, with birds singing on the logical processes (3)—reveal scale invari- tion is therefore usually attributed to QTLs ance, suggesting that simple rules underlie the of large effect. structure and function of ecosystems. Some D. Storch is at the Center for Theoretical Study, The recent studies of selected maize and Charles University, 110 00-CZ Praha 1, Czech recent discoveries have shed light on the broiler lines (3, 6) were extensive, and the Republic. P. A. Marquet is at the Center for Advanced processes that define quantitative patterns in QTL effects identified were small. These Studies in Ecology and Biodiversity, Departamento de biodiversity. Dissecting these patterns was the appear to conform to the infinitesimal model Ecología, Pontificia Universidad Católica de Chile, goal of an international workshop held Casilla 114-D, Santiago, Chile. K. J. Gaston is in the of genes of small effect assumed in much Biodiversity and Macroecology Group, Department of recently in Prague and co-organized by the quantitative genetic theory (4), which pre- Animal and Plant Sciences, , Santa Fe Institute and the Center for dicts the observed continuous steady Sheffield S10 2TN, UK. Theoretical Study at Charles University (4).

684 4 FEBRUARY 2005 VOL 307 SCIENCE www.sciencemag.org P ERSPECTIVES One major challenge is to relate patterns tional to productivity, accurately predicts the force for the current distribution of biodi- of species richness to the spatial distribu- increase in bird species richness with increas- versity on Earth. tion of individual species. Scale invariance ing environmental productivity. Thus, This “zero sum” rule has been assumed in of the species-area relationship, for exam- species richness is influenced by available neutral theories of biodiversity that attempt ple, has led to the development of models energy in different ways. It is possible that to explain major phenomena concerning relating this phenomenon to the fractal spa- the diversity of ectotherms may principally species richness, abundance, and distribu- tial distribution of individuals, which is be driven by temperature, whereas resource tion in terms of stochastic population characterized by similar spatial patterns availability may be more important for growth, migration, and speciation. The orig- over several scales of resolution (5, 6). But endothermic animals that are able to regu- inal neutral theory formulated by Stephen two questions arise. First, do species really late their body temperature. Hubbell (10) can be extended to incorporate exhibit fractal distributions? Using sophis- The extent to which external constraints realistic dispersal processes (Luis Borda-de- ticated analytical techniques applied at dif- limit biodiversity has long been a topic of Água, University of Georgia) and can predict ferent scales of observation, Jack Lennon debate. Typically, those models relating the many biodiversity patterns—such as the (Macaulay Institute, Aberdeen) and species-area relationship to the distribution relationship between the richness of native Fangliang He (University of Alberta) con- patterns of individual species ignore inter- and alien species (Tomasv Herben, Charles cluded that some species really do, but oth- specific interactions or limits to the number University, Prague). However, current neu- ers do not, and that the level of fractality is of species or individuals that can co-occur tral models are insufficient to explain related in part to the rarity of the species. in an area. But every ecosystem must have observed patterns of species’ distributions.

Second, it is not clear why species’v distri- some finite capacity in terms of the number Jerome Chave (Université ) butions should be fractal. Arnostv Sizling of individuals or biomass that it can sustain. reported that both stochastic spatial (Center for Theoretical Study, Prague) James Brown (University of New Mexico) processes and environmental heterogeneity reported that a distribution that is indistin- pointed out that this leads to a zero-sum sit- (see the figure) contribute to the species dis- guishable from a fractal distribution may uation that may affect biodiversity dynam- tribution patterns of neotropical trees. At emerge from random processes of spatial ics as well as species distribution. least at small spatial scales, the way that aggregation on several scales of resolution. Relationships between environmental species divide resources and habitat is cru- These findings suggest that fractals are sim- energy and biodiversity imply that such cial for understanding patterns of biodiver- ply a useful and analyti- limitations do exist, as sity (Mark Ritchie, Syracuse University). cally tractable approxima- Pure niche-assembly models demonstrated by David That said, it is sometimes necessary to sac- tion of the complex spatial Currie (University of rifice knowledge about the biological peculi- aggregation of individuals Ottawa). He emphasized arities of species and communities to make that is responsible for pat- that large-scale spatial models and theories tractable. Neutral theo- terns of biodiversity. variability of species ries, although unrealistic, provide insight into Another widespread richness is very well the general processes that govern species rich- observation is a positive explained by climatic ness and distribution. John Harte (University relationship between spe- variables, making it of California, Berkeley) demonstrated that cies richness and available unlikely that historical even simple static models without the addition energy (7). Andrew Clarke events were the driving of specific parameters could be useful. His (British Antarctic Survey, model predicts a variety of Cambridge) noted that Pure dispersal-assembly models patterns in the spatial dis- understanding the connection between spe- Biodiversity, species distri- tribution and abundance of cies richness and energy has been hindered bution and community plant species just on the by ignoring the different meanings of the composition. The spatial basis of the frequency dis- word “energy” implied in the variety of structure of species’ distribu- tribution of total species mechanisms invoked as explanations. One tions is driven by the availabil- abundance and a simple important measure of energy is temperature, ity of suitable habitats and of probability rule that relates which presumably influences species rich- individuals that can poten- occupancy at one spatial ness by increasing the rates of a variety of tially colonize a site. In niche- scale to occupancy at biological processes, leading to accelerated assembly models of commu- smaller scales. Although speciation (8). Although we do not yet have a nity structure (above), suit- its biological interpreta- comprehensive theory (9), Andrew Allen able habitat is the major force tion is unclear, this ap- (University of New Mexico) suggested that driving species’ distributions proach shows that different the richness of ectothermic species (which (background color refers to patterns are intrinsically cannot regulate their body temperature) habitat suitability for species linked to one another, and could be predicted from environmental tem- with respective colors). The same pattern of it is important to study these inevitable links, perature according to fundamental laws con- species’distributions and local community compo- for they can provide the basis for understand- sition (right) can, however, be interpreted in terms necting body size, temperature, and meta- ing the dynamics of complex ecological net- of the spatial distribution of sources of potential bolic rate. Another measure of energy is envi- colonists of different species (colored polynoms) works (Neo Martinez, Pacific Ecoinformatics ronmental productivity, which affects the without the need to consider habitat suitability. and Computational Ecology Lab, Berkeley). amount of resources available to a popula- These “dispersal-assembly models” give a reason- As physicist Murray Gell-Mann (Santa tion, enabling an increase in the numbers of able prediction of general properties of observed Fe Institute) observed, it is interesting how individuals and the persistence of more spatial species’distributions and community struc- little we actually know. Although sophisti- species. David Storch (Center for Theoretical ture, but fail to predict details about the local cated techniques of data analysis are now Study, Prague) showed how a simple model species composition of communities. [Adapted available and theory is developing rapidly, of spatial dynamics, which assumes that the with permission from a figure by Jerome Chave, macroecology as an exact science is still in probability of species occurrence is propor- Université Paul Sabatier] its infancy. Many patterns have been docu-

www.sciencemag.org SCIENCE VOL 307 4 FEBRUARY 2005 685 P ERSPECTIVES

mented using only limited data sets, and across multiple scales and bridging the gap 3. J. H. Brown et al., Ecology 85, 1771 (2004). even some phenomena assumed to be well between theory and observation is crucial to 4. Scaling Biodiversity, 19–23 October 2004, Prague, documented (albeit with a multitude of achieve a better understanding of quantita- Czech Republic. 5. J. Harte et al., Science 284, 334 (1999). v potential explanations) may look different tive patterns in biodiversity. However, one 6. A. L.Sizling, D. Storch, Ecol. Lett. 7, 60 (2004). when examined from another perspective. thing is certain: Darwin’s entangled bank is 7. D. Currie, Am. Nat. 137, 27 (1991). For some taxa (such as the bacteria or pro- far more entangled than even he thought. 8. A. P.Allen et al., Science 297, 1545 (2002). 9. D. Storch, Science 299, 346 (2003). tists), we have only weak evidence about 10. S. P. Hubbell, The Unified Neutral Theory of References basic patterns (Brendan Bohannan, Stanford Biodiversity and Biogeography (Princeton Univ. Press, 1. C. Darwin, On the Origin of Species by Means of University; Jessica Green, University of Natural Selection (John Murray, London, 1959). Princeton, NJ, 2001). California, Merced). Obtaining better data 2. K. J. Gaston, Nature 405, 220 (2000). 10.1126/science.1106935

ASTRONOMY nearly 1000 times the field strength in the galactic disk. In addition, the early studies At the Heart of the Milky Way (4) suggested that all filaments are essen- tially perpendicular to the galactic plane T. Joseph W. Lazio and Theodore N. LaRosa (within 20°). The picture that emerged was of a strong, dipolar magnetic field filling t a distance of just 25,000 light years referred to as streaks, threads, and fila- the galactic center. The filaments were (2.5 × 1020 m), the center of our ments. The relation between these isolated explained as magnetic flux tubes, which Agalaxy, the Milky Way, provides the filaments and the bundled filaments of the were “lit up” by relativistic electrons that foundation for understanding phenomena radio arc remains unknown. were accelerated by a local interaction such in other galaxies. The central black hole (1) These filamentary structures are distin- as magnetic field reconnection (4). and regions of intense star formation in its guished by extreme length-to-width ratios However, recent radio and submillime- vicinity can be probed at 100 times the res- (~10 to 100), nonthermal spectra, and a ter observations are challenging this simple olution of even the nearest galaxies. high intrinsic polarization (~30%, and in picture. New wide-field images at radio Nonetheless, even the basic properties of a some cases approaching the theoretical wavelengths (between 20 and 90 cm, see key component of the galactic center, its maximum of 70% for synchrotron radia- the figure) have substantially increased the magnetic field, remain poorly understood. tion). The polarization and nonthermal number of known filaments and have Magnetic fields have the potential to spectra are consistent with the filaments shown that the volume over which fila- transform, store, and explosively release being produced by synchrotron radiation ments occur is much larger than originally energy, to transport angular momentum, from relativistic electrons spiraling around thought. With the larger number of fila- and to confine high-energy plasmas into a magnetic field. Detailed measurements of ments has come the discovery of filaments powerful jet flows. They are therefore cen- individual filaments have shown that the tral to astrophysical activity from stellar to magnetic fields are aligned longitudinally galactic scales. with the filament. Magnetic fields are found throughout Estimates of the field strengths within the Milky Way. Measurements suggest that the filaments are on the order of 0.1 µT— the magnetic field in the spiral disk of our galaxy has two components, one globally ordered and the other random, with approx- –28.40 imately equal strengths of ~0.3 nT (2); the globally ordered component generally fol- lows the spiral arms of the galaxy. Key 50 questions about the magnetic field in the galactic center are whether it is comparable in strength or much stronger than the field –29.00 Magnetic filaments in the galactic in the disk, and whether it is globally center. Adapted from (15). (Left) ordered or largely random. This 330-MHz image acquired at the About 20 years ago, the first high-reso- 10 Very Large Array, shows the central lution radio images of the galactic center region of the Milky Way.The galactic (3) revealed numerous magnetic structures center is located within the burned- that are unique to the galactic center. The out region just above the center of 20 the image, and the plane of the Milky most striking of these is the galactic center (J2000) Declination radio arc, a series of parallel linear fila- Way’s spiral disk runs approximately diagonally through the image. ments, each of which is merely a few light 30 Clearly visible near the top left is the years wide yet more than 100 light years galactic center radio arc, the bundle long. Also observed were a number of iso- of filaments, and, just above it, a lated linear features that were variously 40 newly discovered system of fila- ments. (Top right) This close-up of the region in the white box shows fil- T. J.W. Lazio is with the Remote Sensing Division, Naval aments with a diverse range of orien- Research Laboratory, Washington, DC 20375, USA. 50 E-mail: [email protected] T. N. LaRosa is in the tations as well as two sets of poten- Department of Biological and Physical Sciences, 17 47 00 46 30 004530 00 44 30 00 43 30 tially crossing or interacting fila- Kennesaw State University, Kennesaw, GA 30144, USA. Right ascension (J2000) ments (indicated by arrows).

686 4 FEBRUARY 2005 VOL 307 SCIENCE www.sciencemag.org P ERSPECTIVES that are not perpendicular to the galactic High-resolution submillimeter polariza- to which they are associated with other plane (5, 6). A range of filament orienta- tion observations have also recently pro- objects in the galactic center. These data tions (see the figure, top right panel) is dif- vided insights into the magnetic field at the may reveal whether the magnetic field in ficult to reconcile with the idea of a strong, galactic center. In contrast to radio meas- the galactic center is a result of a dynamo pervasive dipolar field. Moreover, the much urements, which probe the magnetic field in generated by the central black hole (11); a larger volume over which filaments are relatively dilute plasmas, submillimeter large-scale outflow driven by past injec- found implies a much larger total energy observations probe the thermal emission tions of energy from the combined actions contained in the magnetic field. from dust grains in cold, dense clouds. If of many supernovae, the central black hole, The diverse orientations and larger than these dust grains rotate in a magnetic field, or both (12, 13); or even the amplification expected volume for the filaments have they will align perpendicular to it. The ther- of a primordial field predating the forma- prompted reconsideration of their origin. mal emission from the grains is then polar- tion the Milky Way (14). One possibility is that they represent local- ized because of the grain orientation. ized, and potentially dynamic, enhance- Large-scale observations show that the References and Notes ments of an otherwise weak magnetic field. magnetic field in molecular clouds at the 1. G. C. Bower et al., Science 304, 704 (2004). For instance, in one model, the filaments galactic center is aligned preferentially in a 2. R. Beck, Space Sci. Rev. 99, 243 (2001). 3. F.Yusef-Zadeh et al., Nature 310, 557 (1984). are magnetic wakes generated when molec- disk-like orientation, much like the mag- 4. M. Morris, E. Serabyn, Annu. Rev. Astron. Astrophys. ular clouds interact with a wind from the netic field in the spiral disk (8). At higher 34, 645 (1996). galactic center, analogous to cometary resolution, the field traced by submillimeter 5. C. C. Lang, K. R. Anantharamaiah, N. E. Kassim, T. J. W. Lazio, Astrophys. J. 521, L41 (1999). plasma tails formed in the solar wind (7). In polarization measurements is more com- 6. T. N. LaRosa, M. E. Nord, T. J. W. Lazio, N. E. Kassim, other models, the filaments form when col- plex, influenced at least in part by the local Astrophys. J. 607, 302 (2004). liding stellar winds fill locally strong mag- dynamics of the dust (9). In some regions, 7. S. N. Shore,T. N. LaRosa, Astrophys. J. 521, 587 (1999). netic flux tubes. Of course, more than one the field directions inferred from these 8. G. Novak et al., Astrophys. J. 583, L83 (2003). 9. D.T. Chuss et al., Astrophys. J. 599, 1116 (2003). model may be correct. The current models measurements are similar to those found in 10. Q. D.Wang et al., Astrophys. J. 581, 1148 (2002). offer intriguing ideas for the origin of the the synchrotron-emitting plasma. 11. S. K. Chakrabarti et al., Nature 368, 434 (1994). filaments, but none can account for the full These observations at different wave- 12. R.A. Chevalier, Astrophys. J. 397, L39 (1992). 13. J. Bland-Hawthorn, M. Cohen, Astrophys. J. 582,246 range of characteristics exhibitted by these lengths remain to be reconciled with each (2003). structures. Clearly, their presence in the other and with suggestions that the fila- 14. B. D. G. Chandran et al., Astrophys. J. 528, 723 (2000). galactic center indicates that it is the only ments are also visible at x-ray wavelengths 15. M. E. Nord et al., Astron. J. 128, 1646 (2004). region in the Milky Way where they can [see for example (10)]. New instruments 16. We thank D. Chuss, N. Kassim, M. Nord, and S. Shore for helpful discussions. Basic research in radio astronomy form, or more generally, that only in the under development should be able to detect at the Naval Research Laboratory is supported by the nuclei of galaxies are the conditions suffi- many more filaments, determine their ori- Office of Naval Research. cient for filaments to form. entations and lengths, and assess the extent 10.1126/science.1100793

SIGNAL TRANSDUCTION Ste12, which is guided by Tec1 to the pro- moters of filamentous growth genes (4). In Signaling Specificity in Yeast addition, MAPKs use different activation mechanisms—for example, Fus3 must Elaine A. Elion, Maosong Qi,Weidong Chen interact with the Ste5 scaffold to be acti- vated, whereas Kss1 activation does not central problem that continues to puz- program (also called invasive growth in hap- have this requirement (1–3). However, zle biologists is how cells translate loids and pseudohyphal development in these levels of control do not explain how Amyriad stimuli into highly specific diploids). Fus3 and Kss1 are both activated the kinases are insulated from erroneous responses. Signaling specificity is compli- in response to mating pheromone by the cross-activation. For example, why doesn’t cated in eukaryotic cells because signal MAPK scaffold protein Ste5; Kss1 also can activation of mating pheromone by Kss1 transduction pathways that respond to differ- be activated by cell surface proteins such as induce a filamentous growth program? ent stimuli and perform distinct functions mucin (Msb2) and other conditions that pro- One level of insulation comes from atten- often share the same pathway components. mote invasive growth [(1–3); see (4) for a uation of nonessential MAPKs under any In haploid budding yeast, for example, the summary of targets in the mating and inva- given condition. During filamentous growth, mitogen-activated protein kinase (MAPK) sive growth pathway]. How is pathway selective repression of Fus3 by phosphatases cascades that regulate both mating and fila- specificity maintained with so much redun- such as Msg5 enforces the filamentous mentous growth share the same three activat- dancy? Two papers in a recent issue of Cell growth program (1). During pheromone stim- ing kinases—Ste20 MAPKKKK or PAK- reveal how the Fus3 and Kss1 MAPK signal- ulation, Fus3 attenuates the activation of Kss1 type kinase, Ste11 MAPKKK, and Ste7 ing pathways are insulated from each other, and influences its strength and duration of sig- MAPKK (which are activated serially)—as ensuring smooth execution of the mating naling (7). Another key level of insulation well as the Ste12 factor (see the program without erroneous activation of the comes from the inhibition of filamentous figure). Two separate MAPKs are activated filamentous growth program (5, 6). growth by Fus3 and the expression of Tec1- by the same Ste7 MAPKK: Fus3, which is Signaling specificity is possible in part dependent genes during pheromone signaling essential for the mating program, and Kss1, because MAPKs choose different sub- (8). The mysterious mechanism of this potent which is essential for the filamentous growth strates. For example, during mating, Fus3 insulation is now revealed by the two Cell phosphorylates the cell cycle regulator Far1, papers (5, 6) and a third study (9). These whereas both Fus3 and Kss1 can phospho- reports show that the Fus3 and Kss1 signaling The authors are in the Department of Biological rylate Ste12 (see the figure). During fila- cascades are insulated from each other Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, MA 02115, USA. E-mail: mentous growth, Kss1 regulates the Tec1 because Fus3 controls degradation of Tec1, [email protected] transcription factor indirectly through the transcription factor essential for activation

www.sciencemag.org SCIENCE VOL 307 4 FEBRUARY 2005 687 P ERSPECTIVES of the filamentous growth conclusions about the identity of the F-box A Mating pheromone program (see the figure). protein that recognizes phosphorylated Tec1 receptor/– Nutrient Multiple stimuli MAPK scaffold/ Mucin limitation The beauty of these stud- and links it to the SCF ubiquitin-ligase com- and ways of Ste5 ies is that they use estab- plex. Chou et al. (5) propose SCFCdc4 (WD40 sensing stimuli lished yeast mutants that class) as their candidate for the following rea- Common are hyperfilamentous as a sons. Tec1 is stable in a cdc4-3 mutant activating kinases result of mutations in Tec1 although it is phosphorylated at residue 273; (Thr273 switched to Fus3-phosphorylated Tec1 interacts with 274 Dual MAPKs Fus3 methionine, and Pro Cdc4 in vitro; and Tec1 cannot be ubiquiti- Kss1 switched to serine). These nated in a cdc4-3 yeast mutant. In their turn, Unique and yeast mutants point the Bao et al. (6) propose an as yet undefined Dia2 overlapping Far1 Ste12 TEC1 Tec1/Ste12 way to discovering the SCF protein (LRR class by homology) on substrate mRNA critical residue in Tec1 the basis of the stabilization of Tec1 in a dia2 specificity that is phosphorylated by mutant that is known to be hyperfilamentous Mating Filamentous growth Fus3, signaling the degra- (10). Although it is possible that multiple SCF Distinct dation of Tec1. Thr273 and complexes regulate Tec1 [and the data in Chou cell fates Pro274 together form part et al. support short-term stabilization of Tec1 of a consensus phospho- in a dia2 mutant as well as in a grr1 mutant rylation site for proline- that is also hyperfilamentous (10)], more B directed MAPKs (5, 6). work is needed to dissect the possible inter- Pheromone The threonine residue at position 273 is one of play between different SCF complexes and multiple sites phosphorylated by Fus3 in vitro between the direct and indirect effects of (5, 6, 9) and in pheromone-treated cells in mutations on Tec1 production. Fus3 Kss1 vivo (5). The proline residue at position 274 is The most important notion gleaned from predicted to be necessary for recognition of these studies is that pathway specificity can SCF-dependent degradation the threonine by a MAPK. However, Tec1 be regulated by the MAPKs downstream of T273 mutants with a methionine at residue 273 or a the MAPK signaling cascades. Previous Tec1 P Tec1/Ste12 serine at residue 274 are resistant to work supports the possibility that MAPK- pheromone-induced degradation and display induced proteolysis of MAPK targets may be almost the same absence of insulation as a a general means of controlling pathway Filamentous growth yeast mutant that completely lacks the FUS3 specificity (11, 12). Such a regulatory device gene (5, 6). These findings argue com- permits flexibility by focusing control on pellingly that down-regulation of the Tec1 specific MAPK outputs while maintaining protein is the major way in which Fus3 insu- others that may be beneficial for the desired lates the mating MAPK cascade from the fil- program. Proteolysis is advantageous No fuss about insulation. (A) The Fus3 and Kss1 MAPKs are activated by the same signaling amentous growth MAPK cascade. In contrast, because it is rapid and irreversible, allowing cascades—that is, they share the common acti- Kss1 does not phosphorylate Tec1 (5, 6, 9), for sharp transitions. Phosphorylation- vating kinases Ste20 MAPKKKK, Ste11 MAPKKK, ensuring that Tec1 remains stable during fila- induced proteolysis of substrates provides a and Ste7 MAPKK. Although Fus3 and Kss1 are mentous growth. Interestingly, the amount of way to get rid of unwanted by-products of both activated by mating-specific inputs that Tec1 degraded is proportional to the concen- activated protein kinases at sites distal from involve the Ste5 scaffold protein (red; tration of the pheromone stimulus (5). This is the place of activation. There are additional pheromone receptor, G protein–MAPK scaffold), consistent with earlier findings by several opportunities for regulating substrate: at the only Kss1 can be activated independently of groups showing that a low concentration of place and time of substrate degradation, and Ste5 by inputs that promote filamentous growth mating pheromone induces production of during various phosphorylation events that (blue; the mucin Msb2 and nutrient deprivation). TEC1 mRNA and invasive growth. Thus, the are required for substrate to be recognized by These two MAPKs have both common and system may have evolved to permit dividing the proteolysis machinery. These different unique substrates. Fus3 phosphorylates the Far1 yeast both to forage by filamentous growth methods of regulation are also applicable to cyclin-dependent kinase inhibitor and the and to search for optimal mating conditions feedback and cross-regulation of signal Tec1/Ste12 transcription factor, two events crit- while cells are primed for mating, and before transduction pathways. Perhaps Nature has ical for mating. Kss1 regulates the activation of Tec1 is degraded. found a simple strategy to take full advantage Tec1/Ste12, which is necessary for filamentous Additional evidence suggests that phos- of redundancies in a system that has been growth. During mating, Kss1 activation phorylated Tec1 may be recognized for designed to be exquisitely sensitive. switches on expression of both mating and fila- destruction by ubiquitin-dependent proteoly- mentous growth genes, including the TEC1 gene sis through the action of an SCF (Skp1- References (depicted by dashed arrows as they are not Cdc53/Cul1–F-box) ubiquitin ligase complex 1. J.Andersson et al., EMBO J. 23, 2564 (2004). essential for mating and interfere with cell cycle (5, 6). Tec1 is ubiquitinated in the presence of 2. S. Maleri et al., Mol. Cell. Biol. 24, 9221 (2004). arrest, which is needed for terminal differentia- 3. P. J. Cullen et al., Genes Dev. 18, 1695 (2004). tion). (B) Fus3 phosphorylation of the Tec1 pro- mating pheromone (5, 6) but not if residue 273 4. J. Zeitleiger et al., Cell 113, 395 (2003). tein on Thr273 leads to ubiquitin-dependent is mutated or Fus3 is absent (6). In addition, 5. S. Chou et al., Cell 119, 981 (2004). Tec1 is completely stabilized in a cdc34-2 6. M. Z. Bao et al., Cell 119, 991 (2004). degradation of Tec1 by a SCF ubiquitin-ligase 7. W. Sabbagh et al., Mol. Cell 8, 683 (2001). SCIENCE complex, neatly blocking entry into the filamen- yeast mutant (which lacks the E2 enzyme) (5, 8. H. D. Madhani et al., Cell 91, 673 (1997). tation program. Thus, the status of the Fus3 6) and a cdc53-1 mutant (6) in the presence of 9. S. Bruckner et al., Curr. Genet. 46, 331 (2004). MAPK determines whether cells mate or enter a pheromone (although it is possible that these 10. S. P. Palecek et al., Genetics 156, 1005 (2000). filamentous growth program. This regulatory pleiotropic mutations could have interfered 11. A. S. Nateri et al., Science 303, 1374 (2004). 12. M. Gao et al., Science 306, 271 (2004). device is not necessary in diploid cells, which do with the activation of Fus3). The Chou et al.

not mate or express Fus3. (5) and Bao et al. (6) groups reach different 10.1126/science.1109500 CREDIT: SUTLIFF/ KATHARINE

688 4 FEBRUARY 2005 VOL 307 SCIENCE www.sciencemag.org BREVIA chambered noncalcareous taxa form a series of deep-branching evolutionary lineages, rep- Simple Foraminifera Flourish at resenting the radiation from which more complex multichambered groups arose (5). the Ocean’s Deepest Point They include the only foraminifera to have invaded freshwater and terrestrial (6) habitats. 1 2 3 Yuko Todo, Hiroshi Kitazato, * Jun Hashimoto, Our results indicate that these primitive groups Andrew J. Gooday4* are also important components of benthic com- munities in the deepest ocean trenches and may Extreme water depths make it very difficult to This assemblage most resembles the fauna be more adaptable than other foraminiferans to sample the bottom of deep-ocean trenches. As a seen at a shallower site in the Marianas Trench extreme pressures. Similar forms are wide- result, almost nothing is known about small (7123 m water depth) (table S1). Here, spread in the abyssal deep sea and abundant at sediment-dwelling organisms (meiofauna) liv- Nodellum- and Resigella-like allogromiids 7800 m water depth in the Atacama Trench (7), ing in these environments, which are among the were also common, albeit less so (45% of the but they are never a dominant faunal compo- most remote on Earth. During a study of west- assemblage) than at the Challenger Deep. nent. The Challenger Deep may have developed ern Pacific trenches (97000 m water depth), we Psammosphaerids, however, were more abun- to its present depth during the past 6 to 9 million discovered abundant foraminifera (shelled pro- dant (37%) and the assemblage more diverse years (8). Its very distinctive foraminiferal fauna tists) living in the Challenger Deep, probably represent the remnants of an the deepest place (10,896 m) in the abyssal assemblage that was able to world ocean. The fauna is dominated adapt to the steady increase in hydro- by morphologically simple species static pressure over this time period. with organic walls. These distinctive taxa seem to be characteristic of the References and Notes deepest ocean depths. 1. Materials and methods are available as supporting material on Science Online. The 0- to 1-cm layer of a 2. A. J. Gooday, J. Foraminifer. Res. 32, sediment core collected with the 384 (2002). KAIKO Remote Operated Vehicle 3. K. Akimoto, M. Hattori, K. Uematsu, C. Kato, Mar. Micropaleontol. 42, 95 (2001). (1), belonging to the Japan Agency 4. J. Hashimoto, Ed., Onboard Report for Marine Earth Science and KR02-13, KAIKO/KAIREI Cruise in the Technology (JAMSTEC), yielded Challenger Deep (JAMSTEC, Yokosuka, Japan, 2002). 432 rose-bengal–stained, and there- 5. J. Pawlowski et al., Proc. Natl. Acad. fore living, benthic foraminifera Sci. U.S.A. 100, 11494 (2003). (È449 per 10 cm2) (Fig. 1). This 6. R. Meisterfeld, M. Holzmann, J. Pawlowski, Protist 152, 185 (2001). population density was similar to 7. A. Sabbatini, C. Morigi, A. Negri, A. J. that found at our shallower (7088 to Gooday, J. Micropaleontol. 21, 131 (2002). 7761 m) trench sites (table S1) and 8. K. Fujioka, K. Okino, T. Kanamatsu, Y. greater than at many abyssal sites Ohara, Geophys. Res. Lett. 29, 1372, 10.1029/2001 GL 013595 (2002). (2). Except for four individuals of Fig. 1. Live (stained) specimens of benthic foraminifera from the Chal- lenger Deep, part of the Izu-Bonin-Mariana Arc trench system. (A)A 9. We thank the captain and crew of the the multichambered agglutinated needle-shaped, organic-walled allogromiid resembling Chitinosiphon. KAIREI for assistance at sea and S. Tsuchida genera Leptohalysis and Reophax, and D. Kim for processing the core sample. (B) Multichambered, organic-walled allogromiids, representing a Supported in part by JAMSTEC, the Japan the assemblage was dominated by similar to Resigella.(C) Spherical, organic-walled allogromiids. (D)A Society for Promotion of Science, the delicate, soft-walled species. Most spherical, agglutinated psammosphaerid. Scale bars, 50 mm. Kaplan Foundation, and the Natural Envi- (85%) of these 428 specimens were ronment Research Council. organic-walled allogromiids, typically brown in (57 species). Samples from two northwest color and with tubular morphologies, sometimes Pacific trenches (Kurile and Japan; 7761 m Supporting Online Material www.sciencemag.org/cgi/content/full/307/5710/689/ subdivided into two or more elongate or globular and 7088 m, respectively) yielded only a few DC1 chambers. These morphotypes resemble the Nodellum-orResigella-like specimens, suggest- Materials and Methods genera Chitinosiphon, Nodellum,andResigella. ing that the dominance of these forms in the Table S1 Other taxa were spherical, organic-walled Marianas Trench (including the Challenger 20 September 2004; accepted 16 November 2004 allogromiids (5.6%) and spherical and flask-like Deep) may reflect a more oligotrophic setting 10.1126/science.1105407 agglutinated species (psammosphaerids and compared with the northwest Pacific, rather saccamminids) (9.6%). Such a high proportion than water depth. An abyssal sample (Station 1 of organic-walled allogromiids is very unusual; no. 64; 5507 m water depth) was more diverse Department of Life and Earth Sciences, School of Science, Shizuoka University, Shizuoka, Japan, and U. in most deep-sea environments, they constitute than any of the trench material. Psammosphaerids Dom Corporation, Tokyo, Japan. 2Institute for Research 5 to 20% of the living assemblage (1). A and species of Lagenammina were most on Earth Evolution, Japan Agency for Marine Earth, previous study (3) of foraminifera from a common, and the Nodellum and Resigella Science and Technology, Natsushimacho 2-15, Yokosuka 237-0061, Japan. 3Faculty of Fisheries, Nagasaki Univer- comparable water depth reported four stained group made up about 10%, a typical propor- 4 (living) individuals belonging to the aggluti- tion at abyssal depths. Studies on metazoan sity, Nagasaki, Japan. DEEPSEAS Benthic Biology Group, Oceanography Centre, Empress Dock, nated genus Lagenammina. Foraminifera with organisms in video records and samples from European Way, Southampton SO143 ZH, UK. calcareous walls cannot exist at this great the Challenger Deep are in progress (4). *To whom correspondence should be addressed. depth because the water is strongly under- Analyses of small subunit ribosomal DNA E-mail: [email protected] (H.K.) and ang@ saturated in calcium carbonate. gene sequences have suggested that single- soton.soc.ac.jp (A.J.G.)

www.sciencemag.org SCIENCE VOL 307 4 FEBRUARY 2005 689 RESEARCH ARTICLES conformation provides insight into the structural basis for cAMP-induced activation Crystal Structure of a Complex of PKA. Molecular architecture. The architec- Between the Catalytic and ture of the RIa(91–244):C complex reveals an extended interface that covers nearly Regulatory (RIa) Subunits of PKA 3000 A˚ 2 (Fig. 1, A and B, and movie S1). Although the C subunit assumes a fully 1 1,2 1,3,4 Choel Kim, Nguyen-Huu Xuong, Susan S. Taylor * closed conformation with Mn2AMP-PNP bound at the active site cleft, it does not The 2.0-angstrom structure of the cyclic adenosine monophosphate (cAMP)– undergo any other major conformational dependent protein kinase (PKA) catalytic subunit bound to a deletion mutant changes as a result of complex formation; of a regulatory subunit (RIa) defines a previously unidentified extended instead, it serves as a stable scaffold for interface. The complex provides a molecular mechanism for inhibition of PKA docking RIa. The docking surface on the C and suggests how cAMP binding leads to activation. The interface defines the subunit is localized almost exclusively to the large lobe of the catalytic subunit as a stable scaffold where Tyr247 in the G large lobe and is overlapping but distinct helix and Trp196 in the phosphorylated activation loop serve as anchor points from the surface used for docking another for binding RIa. These residues compete with cAMP for the phosphate binding PKA inhibitor, the heat stable protein kinase cassette in RIa. In contrast to the catalytic subunit, RIa undergoes major con- inhibitor (PKI) (Fig. 2D) (15). The binding formational changes when the complex is compared with cAMP-bound RIa.The surface extends from the inhibitor binding inhibitor sequence docks to the active site, whereas the linker, also disordered site at the active site cleft (site 1), across the in free RIa, folds across the extended interface. The b barrel of cAMP binding G helix (site 2) and through to the activation domain A, which is the docking site for cAMP, remains largely intact in the loop (site 3). The electrostatic profile of this complex, whereas the helical subdomain undergoes major reorganization. complex interface is shown in Fig. 1C. In contrast to the C subunit, RIa under- The discovery of cAMP as a second mes- kinase anchoring proteins (7–9). At the C ter- goes major conformational changes upon senger (1) led eventually to the identification minus of each protomer are two tandem complex formation (Fig. 1D and movie S2). of cAMP-dependent PKA (2) with regulatory cAMP-binding domains (CBD-A and CBD- There are three general features that describe subunits that are the major receptors for B), each representing a structural motif that the binding of RIa to form the holoenzyme cAMP (3). PKA, ubiquitous in mammalian has been conserved as a binding module for complex. First, the inhibitor sequence docks cells, regulates processes as diverse as growth, cAMP from bacteria to man (10). Each CBD to the active site cleft. Second, the linker seg- development, memory, metabolism, gene ex- is composed of a helical subdomain and an ment that connects the inhibitor peptide to pression, and lipolysis. The PKA holoenzyme eight-stranded b barrel where cAMP binds. CBD-A is ordered. Third, CBD-A docks onto exists as an inactive complex of two cataytic The essential feature of the b barrel is a the large lobe of the C subunit (Fig. 1C). The (C) subunits and a regulatory (R) subunit conserved phosphate binding cassette (PBC) inhibitor peptide and linker region (Arg94R to dimer; binding of cAMP facilitates dissocia- that anchors the cAMP and shields it from Val112R) are disordered in the crystal struc- tion and activation of the C subunits. solvent (10). The CBDs are joined to the ture of cAMP-bound RIa(91–379), whereas The C subunit is comprised of a small and dimerization domain by a flexible linker (11), in the complex this segment binds as an ex- large lobe with the active site forming a cleft which includes a substratelike inhibitor se- tended chain along the surface of the active between the two lobes. This fold, first quence that docks to the active site cleft of site cleft (Fig. 1, A to D). In addition to this described for PKA, defines the protein kinase the C subunit in the absence of cAMP (12). transition from disorder to order, docking of superfamily (4). The small lobe provides the Whereas structures of dimerization do- RIa results in global reorganization of the binding site for adenosine 5¶-triphosphate mains and the CBDs have been solved by helical subdomain of CBD-A (Fig. 1D). As (ATP), and the large lobe provides catalytic nuclear magnetic resonance (7, 9)andcrystal- predicted by mutagenesis and / residues and a docking surface for peptide/ lography (13, 14), respectively, the structure of deuterium exchange (16–18), this docking site protein substrates. Opening and closing of the an R:C complex has been elusive. Here, we involves the B helix; however, the details and active site cleft is an essential part of report the crystal structure of a holoenzyme magnitude of this conformational reorganiza- catalysis (5). The activation loop in the large complex between an oxidation-resistant form tion were not anticipated. The general confor- lobe contains a phosphorylation site, Thr197, of the C subunit and a deletion mutant of RIa, mation of the b barrel, which provides the essential for catalysis (6). RIa(91–244), which contains the inhibitor se- binding site for cAMP in free RIa,remains Like many signaling proteins, the R quence and the N-terminal cAMP binding do- largely unchanged when the C subunit binds subunits are modular. They contain well- main (CBD-A). The complex was crystallized (Fig. 1D). ordered domains as well as disordered in the presence of a nonhydrolyzable analog of Extended binding surface on the cata- regions. The dimerization domain at the N ATP, adenylyl-imidodiphosphate (AMP-PNP). lytic subunit. Figure 2A shows the ex- terminus is a well-organized four-helix bundle The structure confirms the intrasteric mecha- tended surface on the C subunit that serves that also provides a docking surface for A nism of regulation, defines the intricate order- as a platform for docking RIa.RIa docks to ing of the linker region at the interface of the at least three distinct subsites on the C sub- 1Department of Chemistry and Biochemistry, 2De- 3 RIa and C subunits, reveals a global reorgani- unit (Fig. 1C). Site 1 is where the basic inhib- partment of Physics and Biology, Howard Hughes Medical Institute, 4Department of Pharmacology, zation of the helical subdomain of RIa,and itor sequence docks to the acidic active site University of California, San Diego, CA 92093, USA. shows the unliganded molecular target that is cleft, an interaction shared by all substrates *To whom correspondence should be addressed. seen by cAMP. The comparison of this struc- and inhibitors of PKA. At site 2, the CBD-A, E-mail: [email protected] ture with structures of RIa in its cAMP-bound specifically the hydrophobic portion of the PBC

690 4 FEBRUARY 2005 VOL 307 SCIENCE www.sciencemag.org R ESEARCH A RTICLES containing Tyr205R, binds to an extensive hy- drophobic surface that surrounds Tyr247C in the G helix. Finally, at site 3, the B/C helix in RIa docks to the activation loop of the C subunit. We consider site 1 first. In this complex, AMP-PNP and RIa lock the C subunit into a fully closed conforma- tion. The region of the linker corresponding to the inhibitor site is typically referred to as the P-3 through Pþ1 site (Arg94-Arg-Gly- Ala-Ile98), where the P site is filled by either Ser or Thr for substrates and Gly or Ala for pseudosubstrates. RIa and PKI are pseudo- substrates. As predicted from chemical modifications and from the structure of a 20-oligomer peptide containing the inhibitor consensus sequence of PKI (IP20) bound to the C subunit (15), this segment docks to the active site cleft in a manner similar to IP20 (Fig. 2B and fig. S3). Arg94R (P-3 site) and Arg95R (P-2 site) link the inhibitor sequence to several regions of the C subunit by inter- acting with Glu127C in the linker that joins the small and large lobes, Glu170C in the catalyt- ic loop, and Glu230C in the F helix (Fig. 2C and movie S3). In this closed conformation, Tyr330C in the C-terminal tail interacts with the glycine-rich loop and hydrogen bonds to the 2¶OH group of the ribose at the P-3 site. In addition, His87C in the C helix is hydrogen bonded to the phosphate on Thr197C in the activation loop. Ile98R binds to the Pþ1 loop. This part of the inhibitor sequence in RIa forms a complementary antiparallel b strand that links the glycine-rich loop and the N ter- minus of the C helix to the Pþ1looptoform a b sheet (Fig. 2C). The detailed interactions are summarized in Fig. 3. In spite of sharing a common inhibitor site, IP20 and RIa use different surfaces of the C subunit to achieve high-affinity bind- ing (Fig. 2D). Whereas the amphipathic helix of IP20 docks to a surface that lies N ter- minal to the inhibitor site, RIa docks to the surface that lies C terminal to the inhibitor site. As seen in Fig. 4A, binding of the Pþ1 residue (Ile98R) to the Pþ1 loop not only provides a local docking site but also appears to nucleate an extended hydrophobic inter- face between the C subunit (site 2) and Fig. 1. Stereoview of the RIa(91–244):C:Mn2AMP-PNP complex. (A) The small lobe (residues 11 to CBD-A of RIa. A key element of this sur- 120) and large lobe (residues 121 to 350) of the C subunit are colored in gray and tan, respectively. face on the C subunit is the solvent exposed The N and C termini (residues 11 to 350) are labeled NC and CC. The activation loop, the G helix Tyr247C in the G helix, which is surrounded (G), and the glycine-rich loop are colored in green. Two phosphorylated residues, Thr197C and 338C by a cluster of hydrophobic residues (Figs. 3 Ser , are shown as red spheres. The N and C termini (residues 91 to 242) of RIa are labeled NR and 4A). Docking of Ile98R to the Pþ1 loop and CR. The inhibitory/linker region (IS) (residues 91 to 112) is colored in red, the PBC (P) in yellow, the B/C helix in gray, and the rest in cyan. AMP-PNP is shown in red. This figure was generated with specifically links this hydrophobic cluster by - 247C Molscript (46) and Raster3D (47). (B) Looking down the C:RIa complex, rotated 90 around a means of Tyr to the hydrophobic surface horizontal axis from the view in (A). (C) Electrostatic surface potential of the complex (left) and surrounding Tyr205R in the PBC of RIa with its interface opened up to view the surfaces of individual subunits (right). The linker segment (Figs. 3 and 4, A and B). In its cAMP-bound complements site 3, the PBC complements site 2, and the inhibitor site complements site 1 of the state, cAMP is firmly anchored to the PBC C subunit. This figure was generated with GRASS (48). (D) Structural comparison of RIa in the through its interactions with two key residues, cAMP-bound and C-bound conformations. The inhibitory/linker region (residues 91 to 112, in red) Arg209R and Glu200R (Fig.4C).Thiscomplex is disordered in the cAMP-bound conformation and ordered in the C-bound conformation. The PBC (residues 199 to 210, in yellow) is stretched away from the b barrel when the C subunit binds. The of RIa and C shows that the C subunit also B and C helices become one extended helix (residues 226 to 242, in gray) in the complex. Tyr205R competes for the PBC by the interaction of (yellow) at the tip of the PBC changes its orientation to interact with Tyr247C at the G helix of the C Tyr247C of C with Tyr205R and Ile98R of RIa subunit. This figure was generated with Molscript (46) and Raster3D (47).

www.sciencemag.org SCIENCE VOL 307 4 FEBRUARY 2005 691 R ESEARCH A RTICLES (Fig. 4B). The tip of the PBC in RIa is clear- (Glu105R), interact with Trp196C and Arg194C highly conserved PBC, embedded in the b- ly stretched by these interactions, and this in the activation loop (Figs. 3 and 5C). barrel subdomain, is the anchoring site for stretched PBC has lost its capacity to bind Mutagenesis predicted that Lys213C in the cAMP, as described earlier. The second ele- cAMP (Fig. 4C). Dislodging cAMP from the C subunit formed an ion pair with Glu143R in ment is the B/C helix (Fig. 6B). This element, PBC is probably the key rate-limiting step RIa (19). The structure confirms this inter- which serves as a docking surface for inter- required for holoenzyme formation. action and, in addition, shows that Lys213C acting with other proteins or domains, is teth- Site 3 on the docking surface of C is brings together the A and C helices, which ered to the PBC by a hydrophobic ‘‘hinge’’ dominated by the activation loop containing are far apart in cAMP-bound RIa (Fig. 5D). between the B helix and the tip of the PBC Trp196C and Arg194C. They both interact with Lys213C is located in a highly conserved seg- (13, 22). This RIa:C complex is the first time Glu105R at the linker segment of RIa (Fig. ment that links the activation loop with the F that any CBD has been seen in both a cAMP- 5A). Earlier mutagenesis (19) and genetic helix; we refer to this segment as the APE-F bound state and in a cAMP-free state bound to (20) studies indicated that Trp196C would linker (Fig. 6A), where Glu208C in the APE another protein. Removing cAMP from this play an important role at the R:C interface. motif provides the conserved C-terminal an- deletion mutant of RIa is not sufficient to in- The phosphate of Thr197C links the C helix chor for the activation loop. Whereas Lys213C duce a major conformational change (23). It by means of His87C to the activation loop, interacts directly with RIa, two other nearby is, rather, binding of the C subunit that induces and both of these segments of the C subunit residues, Tyr215C and Lys217C, reach up and major conformational changes (Fig. 6C). interact directly with RIa. His87C also in- buttress the activation loop by hydrogen Appreciation of RIa(91–244) in its cAMP- teracts with Gln84C, and Gln84C in turn binds bonding to the backbone carbonyls of Thr197C bound state compared with its conformation in to Ser99R in the linker region of RIa (Fig. and Ser191C, respectively. A peptide corre- the complex can be best achieved by focusing 5A). Thus, like the glycine-rich loop, the N sponding to this region was also predicted to on three regions (Figs. 1D and 6C). We dis- terminus of the C helix is an integral part of be part of the RIa:C interface based on its cussed many specific residues earlier, and here the RIa:C interface. The phosphate on protection from deuterium exchange (17, 18). we consider these global changes in RIa.The Thr197C is also anchored to the catalytic loop Interaction sites on RIa. Based on a charged and hydrophilic linker region, com- at the active site through its interactions with comparison of six cAMP-bound structures of pletely disordered in cAMP-bound RIa,now Arg165C (Fig. 5B and fig. S2). Two regions CBDs, two elements of the CBD are pre- becomes ordered at the interface (fig. SA). of RIa, the C helix (Met234R) and the linker dicted to be essential for signaling (21). The The B and C helices snap into a single fully

Fig. 2. Surfaces of the C subunit that inter- act with RIa.(A) The complex interface on the large lobe with the F and G helices highlighted in gray. The activation seg- ment (residues 184 to 208) is yellow. Asp208C and Arg280C are each indicated by a ball. Phe239C (49)is part of the hydropho- bic docking site for PKI. Site 1 corre- sponds to the inhibi- tor docking site, site 2 to the G helix with Tyr247C, and site 3 to the activation loop containing Arg194C and Trp196C. This figure was generated with Insight II (50). (B) The inhibitor/linker segment of RIa(91–112). Structure alignment of inhibitor consensus sequences of PKI (inhibitor peptide IP20: Protein Data Bank accession code 1ATP) and RIa. of RIa are colored in gray. The consensus sequence and the amphipa- thetic helix of IP are shown as red bond and ribbon, respectively. Similar to IP20 (Arg-Arg-Asn-Ala-Ile), the in the C-terminal tail, and AMP-PNP. Arg95R at the P-2 site is tightly held inhibitory sequence (Arg94-Arg-Gly-Ala-Ile98)ofRIa blocks the active by Glu170C in the catalytic loop and Glu230C in the F helix (Fig. 3). Farther site of the C subunit. The coordinates of these residues are nearly down toward the linker, Ile98R and Ala100R make antiparallel b-sheet–like identical with an exception of Gly/Asn substitution at the P-1 site. The interactions with the Pþ1 loop. This figure was generated with Visual linker segment interacts not only with the C subunit but with the B/C Molecular Dynamics (VMD) (51) and Molscript (46). (D) Surfaces of the C helix. Specific interacting residues from C and RIa are indicated in black subunit used for recognition of PKI and RIa. This surface rendering of the and red, respectively (arrows). As in the IP20 complex, the C subunit C subunit shows the small lobe in white and the large lobe (residues 128 within the complex is in the closed conformation, with a root mean to 350) in tan. The inhibitors of C, PKI, and the R subunits use a common square deviation of 0.42 A˚2 for equivalent Ca atoms. This figure was surface (magenta) on which a pseudosubstrate (PKI and RI) or a substrate generated with Insight II (50). (C) The inhibitor region (residues 91 to (RII) docks to the active site cleft. To achieve high-affinity binding re- 100) bound to the active site cleft (site 1). The R- and C-subunit residues quires an additional peripheral recognition site (PRS). PKI uses the sur- are drawn with gray and tan carbons, respectively. Arg94R at the P-3 site face shown in blue, designated as PBS 1, whereas RIa uses the surface interacts with Thr51C in the glycine-rich loop, Glu127C in the linker, Tyr330C shown in red (PRS2). This figure was generated with VMD (51).

692 4 FEBRUARY 2005 VOL 307 SCIENCE www.sciencemag.org R ESEARCH A RTICLES extended helix where residues that were pre- In contrast, the helical subdomain of CBD-A ties. Although all share a common substrate- viously fully exposed to solvent are now em- undergoes major conformational changes as like inhibitor site that binds to the active site bedded at the RIa:C interface. Finally, the it docks to the surface of the C subunit. Most cleft, the peripheral sites that allow each to remainder of the helical subdomain that in- notably, the B and C helices become a single bind with high affinity are distinct (Fig. 2, B to cludes the X:N helix and the A helix together extended helix (Figs. 1D and 6D). Trp196C in D, and fig. S1). PKI uses a rather small surface with the interlinking loop rearranges to fill the the activation loop of the C subunit helps to for docking an amphipathic helix that is N space that was vacated by the movement of anchor this extended helix through its inter- terminal to the inhibitor site (15), whereas RIa the C helix (Figs. 1D and 6C). actions with Met234R in the C helix of RIa uses a very large surface C terminal to the As described earlier, docking of the (Figs. 3 and 5C). In its cAMP-bound confor- inhibitor site and a mechanism that involves inhibitor site to the active site cleft results mation, this entire surface of the B/C helix major conformational changes in the helical in ordering of the remaining linker. This of the CBD-A is exposed to solvent (Fig. 1D). region of RIa (Fig. 2D). Preliminary evidence, region, highly conserved in RIa, contributes This surface, which includes Arg226R,Arg230R, based on mutagenesis and hydrogen-deuterium many charged residues (Figs. 1C and 2C). Arg231R,Met234R,andLys240R, now becomes exchange, suggests that RIIb may use yet By interacting not only with C but also with embedded at the interface created by the another variation (26). Collectively, these sites the CBD, they function almost like a zipper large lobe of the C subunit (Fig. 5C). span a substantial surface of the large lobe and to fasten the CBD-A to the large lobe of the Mechanism of inhibition and activa- define the large lobe as a major scaffold. In C subunit. Most of the conformational changes tion. Understanding how a protein kinase is spite of the extended surface, the docking in CBD-A are in the helical subdomain, with inhibited is as important as understanding how sites shown in Fig. 2A are all preformed; no the exception of the tip of the PBC where it functions as a catalyst. Whereas apprecia- conformational changes are induced in the C cAMP docks. This segment is extended tion of the kinase as a catalyst has focused subunit, other than the closing of the active slightly by its interactions with Tyr247C in primarily on the dynamic small lobe and the site cleft, as a consequence of these inhibitors C and Ile98R in RIa (Figs. 1D, 4C, and 6C). opening and closing of the active site cleft (5), binding. PKI, like RIa, requires two Mg2þ The rest of the b barrel remains unchanged. this structure of the RIa:C complex draws our and ATP to form a stable complex (27), attention to the large lobe and reveals the and the C subunit must assume a fully closed importance of the C subunit as a scaffold. conformation. Formation of holoenzyme with PKA has at least seven known physiological RII subunits does not require ATP. inhibitors, four functionally nonredundant R- In addition to the inhibitor site, both the subunit isoforms (24), and three PKI isoforms G helix and activation loop of the large lobe (25); all bind to C with subnanomolar affini- are essential for binding to RIa. In particular,

Fig. 4. Detailed features of site 2. (A) Extended hydrophobic interface created by binding of the Pþ1 residue (Ile98R) to the active site. On the left is the VDW surface of the C subunit and on the right is the complementing surface of RIa, both shown with the inhibitor/linker re- gion and B/C helix of RIa. At the center are two surfaces together as they exist in the complex. Surfaces of RIa and C subunits are colored in yellow and gray, respectively. The inhibitor/linker region, shown as a ribbon, is color coded as follows: inhibitor sequence (residues 94 to 98), tan; linker segment (residues 99 to 105), green; residues 106 to 113, cyan. In cAMP-bound RIa, this entire segment is disordered. B and C helices are also in cyan. The surface of the C subunit is composed of Pþ1 loop and the hydrophobic face of the G helix. Two elements of the surface within the RIa are the hydrophobic residues from the

helical tip of PBC and the X:NHelix to AHelix loop. The surface of the C subunit appears to be preformed, whereas the two elements forming the hydrophobic surface of RIa are brought together by conformational changes. (B) cAMP-binding site in the holoenzyme complex. Tyr205R at Fig. 3. Specific interactions between the R and PBC forms a hydrogen bond (dotted line) with Tyr247C in the G helix. Surrounding this head-on C subunits. The location of the each residue hydrogen bonding between two tyrosines are clusters of hydrophobic residues from both subunits, within the complex is listed alongside of each as shown in (A). The side chain of the Pþ1 Ile nucleates this site. (C) The PBC (residues 199 to 210) amino acid. Solvent molecules at the interface of RIa in the cAMP-bound and C-bound conformations are aligned. The side chains, as in the C- are shown as W. Ion pair, hydrogen-bond, and bound conformation, are yellow, and the cAMP-bound conformations are gray. In addition to van der Waals interactions are notated as @Y, Tyr205R, Asp170R—which orients an important cAMP-binding residue, Arg209R—changes its rotomer Y, and &*&, respectively. position when C binds. All of the images were generated with VMD (51).

www.sciencemag.org SCIENCE VOL 307 4 FEBRUARY 2005 693 R ESEARCH A RTICLES

Fig. 5. Specific interactions associated with site 3. (A) Hydrogen-bonding (50). (C) Interactions between the activation loop and the B/C helix. The network at site 3. Carbons from RIa are gray; carbons from the C subunit interface of Trp196C/Arg194C with the B/C helix of RIa is shown. Specific are tan. Specific interactions are also summarized in Fig. 3. (B) Interactions interactions between Trp196C/Arg194C of the C subunit and Glu105R/Met234R of the activation loop. Phosphorylation of Thr197C creates parts of the of RIa are highlighted. (D)Glu143R in the A helix binds to Lys213C of the RIa:C interface. Each plays a critical role. Whereas Leu198C, Thr201C, and C subunit (Fig. 3) as predicted by mutation study (16). The C helix and Gly200C contribute to recognition of the peptide, Trp196C and Arg194C the A helix of RIa interact closely, thus helping to stabilize the inhibitory provide a critical phosphorylation-dependent docking site for RIa. conformation. All of the images except (B) were generated with VMD Asterisk is the reactive cystine. This figure was generated with Insight II (51).

Tyr247C in the G helix and Trp196C in the ac- this element as part of the RIa:C interface for catalysis (6, 39, 40). The RIa:C structure tivation loop are important key residues that (18). We predict that the G helix will be a highlights another important consequence of anchor RIa (Figs. 4, A and B, and 5C). Be- dynamic and solvent-accessible sensor for pro- phosphorylating the activation loop: creating a cause both are highly conserved as aromatic tein docking in many protein kinases. binding surface for RIa.Theimportanceof residues in the protein kinase family, protein The importance of a single phosphate in Trp196C for R-subunit inhibition was first rec- recognition will likely be a common feature the activation loop was appreciated immedi- ognized by a genetic screen designed to iden- shared by these residues in many protein ki- ately when the first set of crystal structures tify mutants that could not be regulated (20), nases (28). The importance of the G helix as a was solved (4, 31–34). In its fully phospho- and we confirmed experimentally that Trp196C hydrophobic docking site was demonstrated rylated and activated state, p-Thr197C is linked was an essential feature of the R:C inter- previously in the crystal structure of cdk2 to the active site through Arg165C (Fig. 5B) face (19). However, this is only true when bound to kinase-associated phosphatase (35). In inactive kinases, such as cdk2 (31), the C subunit is phosphorylated on Thr197C; (KAP), the phosphatase that removes the the loop is not phosphorylated and is typically dephosphorylated C subunit cannot bind RIa activation loop phosphate from Thr160 (29). disordered. The essential phosphorylation site (41). As described earlier, Trp196C is packed In that complex, the phosphate on Thr160 of on the activation loop is always coupled with against residues in the B/C helix of RIa(91– cdk2 binds directly to the catalytic site of an Arg-Asp (RD) motif (Arg165-Asp166 in the 244), and, similar to Tyr205R, these residues are KAP. Unlike the E, F, and H helices, which C subunit), in which the equivalent to Arg165C fully exposed to solvent when cAMP is form the highly stable core of the large lobe is anchored to the phosphate with Asp166C bound to D(1–91)RIa (Figs. 5C and 6B). and are extremely resistant to deuterium positioned at the active site in the catalytic The most notable revelation that emerges exchange (30), the G helix is more solvent loop, where Asp166C functions as a catalytic base from the RIa:C complex is the remarkable accessible. Its backbone amides are only andprotonsink(36–38). Biochemical studies conformational malleability of RIa. Clearly, shielded from solvent when inhibitors are with mutant proteins have demonstrated the im- the extended linker region needed to become bound. This protection enabled us to identify portance of the activation loop phosphorylation ordered upon binding to the C subunit, and it

694 4 FEBRUARY 2005 VOL 307 SCIENCE www.sciencemag.org R ESEARCH A RTICLES Fig. 6. (A) The APE-F linker motif anchors the activation loop. The activation segment and the APE-F linker motif are rendered as dark gray and light gray ribbons, respec- tively. (B) Conserved features of the CBD in RIa. CBD-A is highly con- served where two charged residues, Arg209R and Glu200R, anchor the cAMP. Hydrophobic residues, shown with the van der Waals surfaces, surround the cAMP and shield the adenine ring. The B and C helices (cyan) interface with the PBC (red) through a hydrophobic hinge at the B helix and through hydrophobic capping of the adenine ring by Trp260R. In our construct, residues 248 to 260 (gold) are missing. This figure was adapted from figure 7C of (21). (C) Comparison of RIa in its cAMP-bound (residues 113 to 242) (14) and C subunit–bound (residues 91 to 242) conformations. The C subunit–bound conformation is on the left and cAMP-bound confor- mation is on right. The inhibitory/ linker region and X:N and A helices are in yellow. PBC is in red and the B and C helices are in cyan. (D) Structure alignment of the two conformations of RIa. Specific conformational changes associated with the B and C helices (residues 226 to 242), the X:N and A helices (residues 113 to 150), and the b barrel (residues 151 to 225) are shown. The C-bound conformation is in cyan and the cAMP-bound conformation is in gray with PBC colored red in both conformations. The b-barrel subdomains minus the PBC (residues 151 to 200 and 210 to 225) of the two conformations align with a Ca root mean square deviation of 1.05 A˚2, whereas the helical subdomains do not align. All of the figures were generated with Insight II (50). was not possible to predict the molecular fea- Ligand-induced activation of the RIa holo- in this complex, thus predict that there will be tures of this docking in advance of the struc- enzyme is a highly cooperative and ordered a major change in the orientation of CBD-B ture. However, the reorganization of the allosteric process with cAMP binding first to relative to CBD-A when RIa is bound to the helical subdomain was not anticipated and CBD-B and then to CBD-A (44). Binding of C subunit. A notable conformational change the magnitude of the conformational changes cAMP to CBD-A then leads to release of the was observed from the neutron scattering is substantial (Figs. 1D and 6C). Although the C subunit. In the complex examined here, studies of the RIa holoenzyme compared to b barrel that provides the docking site for the CBD-B is missing. This domain is tightly an- the RIa homodimer, and this data can now phosphate of cAMP remains largely intact, chored by multiple hydrophobic interactions be modeled more precisely with the avail- the helical subdomain is completely reorga- to the segment that follows the C helix (resi- ability of this RIa:C complex (45). Another nized. Understanding the dynamics and the dues 245 to 259). Trp260R at the beginning of challenge now will be to confirm the spatial pathway for this change is another challenge the A helix of CBD-B provides the hydro- and temporal features of these global changes for the future. We predict that in the absence phobic cap for the adenine ring of cAMP in conformation in the larger protein. of cAMP, the helical subdomain should be bound to CBD-A (Fig. 6B). Thus, when cAMP quite dynamic. The molecule is much less is released from the CBD-A, it removes a References and Notes stable, and unfolding is no longer cooperative major hydrophobic anchor between the two 1. E. W. Sutherland, T. W. Rall, J. Biol. Chem. 232, 1077 when cAMP is removed (42, 43). A stable domains, as predicted by Berman et al.(21). (1958). 2. D. A. Walsh, J. P. Perkins, E. G. Krebs, J. Biol. Chem. conformation is created only in the presence Because CBD-B is tightly anchored to resi- 243, 3763 (1968). of cAMP or C subunit. This mechanism of dues 245 to 259 (14), it will move along with 3. G. N. Gill, L. D. Garren, Proc. Natl. Acad. Sci. U.S.A. stabilizing very different conformations by the C helix of CBD-A when the C subunit is 68, 786 (1971). 4. D. R. Knighton et al., Science 253, 407 (1991). either a small molecule ligand or another bound. The conformational changes in the heli- 5. D. A. Johnson, P. Akamine, E. Radzio-Andzelm, protein has not been observed previously. cal subdomain of CBD-A in RIa, as revealed Madhusudan, S. S. Taylor, Chem. Rev. 101, 2243 (2001).

www.sciencemag.org SCIENCE VOL 307 4 FEBRUARY 2005 695 R ESEARCH A RTICLES

6. J. A. Adams, M. L. McGlone, R. M. Gibson, S. S. Taylor, 29. H. Song et al., Mol. Cell 7, 615 (2001). 52. We would like to dedicate this article to Dr. Shmuel Biochemistry 34, 2447 (1995). 30. S. M. G. Jie Yang, Mike S. Deal, Ganesh S. Anand, Shaltiel formerly of the Weizmann Institute, whose 7. P. Banky et al., J. Mol. Biol. 330, 1117 (2003). Virgil L. Woods, S. S. Taylor, J. Mol. Biol. (2004). encouragement over the past decade helped to drive 8. M. G. Newlon et al., EMBO J. 20, 1651 (2001). 31. H. L. De Bondt et al., Nature 363, 595 (1993). this work. The structure validates his many chemical 9. M. G. Newlon et al., Nature Struct. Biol. 6, 222 (1999). 32.S.S.Taylor,E.Radzio-Andzelm,Structure 2, 345 (1994). insights. We thank N. Nguyen at UCSD x-ray facility 10. J. M. Canaves, S. S. Taylor, J. Mol. Evol. 54, 17 (2002). 33. F. Zhang, A. Strand, D. Robbins, M. H. Cobb, E. J. for his assistance with data collection; C. Ralston, 11. J. A. Hauer, P. Barthe, S. S. Taylor, J. Parello, A. Padille, Goldsmith, Nature 367, 704 (1994). G. McDermott, and the Advanced Light Source (ALS) Protein Sci. 8, 545 (1999). 34. D. J. Owen, M. E. Noble, E. F. Garman, A. C. staffs for their assistance with data collection at ALS 12. J. D. Corbin et al., J. Biol. Chem. 253, 3997 (1978). Papageorgiou, L. N. Johnson, Structure 3, 467 (1995). beamline 8.3.1 (Lawrence Berkeley National Laborato- 13.T.C.Diller,N.H.Xuong,S.S.Taylor,Structure 9, 73 (2001). 35. C. Venien-Bryan et al., Structure 10, 33 (2002). ry); M. Deal for the C subunit; S. Brown for assistance 14. Y. Su et al., Science 269, 807 (1995). 36. F. Li et al., J. Mol. Biol. 315, 459 (2002). with the R subunit; G. Anand for insightful discussion 15. D. R. Knighton et al., Science 253, 414 (1991). 37. B. Nolen, S. S. Taylor, G. Ghosh, Mol. Cell 15, 661 (2004). of the complex formation; S. Hsu and A. Tran for 16. R. M. Gibson, Y. Ji-Buechler, S. S. Taylor, J. Biol. 38. M. Valiev, R. Kawai, J. A. Adams, J. H. Weare, J. Am. technical support; D. A. Johnson (UC Riverside) for Chem. 272, 16343 (1997). Chem. Soc. 125, 9926 (2003). critical reviewing of the manuscript; and N. Haste 17.G.S.Anand,C.A.Hughes,J.M.Jones,S.S.Taylor, 39. J. A. Adams, Chem. Rev. 101, 2271 (2001). with Elzbieta Radzio-Andzelm for assistance with E. A. Komives, J. Mol. Biol. 323, 377 (2002). 40. L. M. Stevenson, M. S. Deal, J. C. Hagopian, J. Lew, figures. This work was supported by NIH grants 18. G. S. Anand et al., Proc. Natl. Acad. Sci. U.S.A. 100, Biochemistry 41, 8528 (2002). GM19301 and GM34921 to S.S.T., Hemoglobin and 13264 (2003). 41. G. H. Iyer, M. J. Moore, S. S. Taylor, J. Biol. Chem., Blood Protein Chemistry Grant, NIH DK07233 to 19.R.M.Gibson,S.S.Taylor,J. Biol. Chem. 272, 31998 (1997). in press. C.K. The Advanced Light Source is supported by the 20. S. A. Orellana, G. S. McKnight, Proc. Natl. Acad. Sci. 42. J. M. Canaves, D. A. Leo´n, S. S. Taylor, Biochemistry Director, Office of Science, Office of Basic Energy U.S.A. 89, 4726 (1992). 39, 15022 (2000). Sciences, Materials Sciences Division, U.S. Department 21. H. M. Berman et al., Proc. Natl. Acad. Sci. U.S.A. 102, 43.D.A.Leo´n, W. R. G. Dostmann, S. S. Taylor, Biochemistry of Energy under contract DE-AC03-76SF00098 at 45 (2005). 30, 3035 (1991). Lawrence Berkeley National Laboratory. Coordinates 22. H. Rehmann et al., Nature Struct. Biol. 10, 26 (2003). 44. D. Øgreid, S. O. Døskeland, FEBS Lett. 129, 287 (1981). for the structure reported above have been deposited 23. C. Kim, unpublished data. 45. W. T. Heller et al., J. Biol. Chem. 279, 19084 (2004). in the Protein Data Bank (accession code 1U7E). 24. E. P. Brandon, R. L. Idzerda, G. S. McKnight, Curr. 46. P. J. Kraulis, J. Appl. Crystallogr. 24, 946 (1991). Supporting Online Material Opin. Neurobiol. 7, 397 (1997). 47. E. A. Merritt, D. J. Bacon, Methods Enzymol. 277,505 www.sciencemag.org/cgi/content/full/307/5710/690/ 25. D. A. Walsh, K. L. Angelos, S. M. Van Patten, D. B. (1997). DC1 Glass, L. P. Garetto, in Peptides and Protein Phospho- 48. M. Nayal, B. C. Hitz, B. Honig, Protein Sci. 8, 676 (1999). Materials and Methods rylation, B. E. Kemp, Ed. (CRC Press, Boca Raton, FL, 49. Single-letter abbreviations for the amino acid resi- Figs. S1 to S3 1990), pp. 43–84. dues are as follows: A, Ala; C, Cys; D, Asp; E, Glu; Table S1 26. D. Law, G. Anand, E. A. Komives, S. S. Taylor, L. F. Ten F, Phe; G, Gly; H, His; I, Ile; K, Lys; L, Leu; M, Met; N, Asn; P, Movies S1 to S3 Eyck, in preparation. Pro;Q,Gln;R,Arg;S,Ser;T,Thr;V,Val;W,Trp;andY,Tyr. References 27. F. W. Herberg, M. L. Doyle, S. Cox, S. S. Taylor, 50. Insight II, Accelrys, San Diego, CA (2003). Biochemistry 38, 6352 (1999). 51. W. Humphrey, A. Dalke, K. Schulten, J. Mol. Graph. 27 August 2004; accepted 1 December 2004 28. S. K. Hanks, T. Hunter, FASEB J. 8, 576 (1995). 14, 33 (1996). 10.1126/science.1104607

angular resolution projected to 3000 km at Saturn’s Temperature Field Saturn_s distance of about 10 astronomical units. Using the LWS 10-by-10–arc sec field from High-Resolution of view, we created mosaics of Saturn using narrow-band filters with effective wavelengths of 8.00, 17.65, 18.75, 23.10, and 24.50 mm. Middle-Infrared Imaging Only at 17.65 mm was the planet successfully G. S. Orton and P. A. Yanamandra-Fisher mosaicked a second time, extending our cov- erage to 270- of longitude.

Saturn was imaged between 8 and 24.5 micrometers at È3000-kilometer The filters chosen are sensitive to CH4 resolution with the Keck I Telescope. Saturn’s atmosphere has zonal tem- emission (8.00 mm) from the 3-mbar region _ perature bands, which are mostly uncorrelated with visible cloud reflectivity, of Saturn s stratosphere and to H2 emission strong 100-millibar zonal temperature oscillations near 32-S, a warm south (17.65 to 24.50 mm) from the 80- to 200-mbar polar cap, and a compact hot point within 3- of the south pole. region of Saturn_s upper troposphere and low- er stratosphere, near the tropopause (fig. S1). Saturn_s atmosphere in the thermal infrared substantial differences in seasonal radiative We used the five filtered observations to derive has hemispherically asymmetric thermal prop- forcing, given that Saturn_s obliquity is 24- temperatures at two levels, near atmospheric erties (1), which indicate the strong influ- as compared with Jupiter_s3-.Butwealso pressures of 3 mbar and 100 mbar, because the ence of seasonal radiative forcing. It also has wanted to document manifestations of dy- contribution functions for the upwelling radi- heterogeneous and time-variable cloud layers namical forces which might be similar to or ance for the 17.65- to 24.50-mm filters, sensed at 5.2 mm(2). These demonstrate the different from Jupiter, such as the degree of clustered around atmospheric levels of 80 to role of dynamics in creating inhomogeneities correlation between Saturn_s thermal and cloud 200 mbar of pressure, were highly overlapping in a cloud system (which is deeper than the fields, the presence of longitudinal temper- and widely separated from the 8.00-mm filter haze layer, which makes its visual appear- ature variations (including waves), and the contribution function peak near 3 mbar (fig. ance so featureless). By imaging at longer appearance of polar vortex phenomena. S2). We derived temperatures at each of these wavelengths that are dominated by the opac- We imaged Saturn at middle-infrared levels, using a model perturbation approach

ity of the well-mixed gases CH4 and H2,we wavelengths during 6 cumulative hours on similartothoseusedtoderivetemperatureson mapped Saturn_s stratospheric and upper the night of 3 February 2004, at the Keck I Jupiter (3, 4). Initial model temperature profiles tropospheric temperature fields in order to Telescope, using the Long Wavelength Spec- were based on Voyager Infrared Interferometer quantify and understand the influences of trometer (LWS), a facility middle-infrared Spectrometer (IRIS) (5–8) and Composite In- dynamics and radiation on the atmosphere camera-spectrometer in its imaging mode. frared Spectrometer (CIRS) (9)results.Tosim- and their similarities with and differences Because angular resolution in the middle in- plify language in this report, we refer to the from the atmosphere of Jupiter. We expected frared is limited by diffraction rather than 3-mbar and the 100-mbar levels as the strato- terrestrial atmospheric turbulence, a large pri- sphere and troposphere, respectively (10). Jet Propulsion Laboratory, MS 169-237, 4800 Oak mary mirror provides unprecedented spatial The primary distinctive feature of the Grove Drive, Pasadena, CA 91109, USA. resolution. Our characteristic 0.5–arc sec stratospheric temperature field (Figs. 1 and 2)

696 4 FEBRUARY 2005 VOL 307 SCIENCE www.sciencemag.org R ESEARCH A RTICLES is its hemispherical asymmetry, with tem- peratures increasing monotonically toward the south pole. This is expected as a result of seasonal forcing, because Saturn is just past the southern summer solstice. Some time- dependent radiative-convective model results (11, 12), shown for this season in Fig. 2, are consistent with the general trend of warming toward higher southern latitudes, but fail to predict the distinctive warming between 70-S (planetocentric) and the pole. A steep temperature increase of È2K takes place at 100 mbar between 69- and 74-S planetocentric latitude, correlated with an in- Fig. 1. Examples of Saturn mosaics at 17.65 mm(left) and 8.00 mm(right), with the appropriate crease of È5 K at 3 mbar (Fig. 2). At 100 mbar, brightness temperatures (Tb) given by the scale bar. Ring obscuration affects the northern hemi- another marked rise of some È2.5 K takes sphere in both images, but they are too faint to be seen against the sky at 8.00 mm. place between È87- and 90-S, with a cor- related increase of È1Kat3mbar.Thesere- gions of elevated temperatures are correlated with lower visible reflectivities. Thus, the vis- ibly dark polar spot is coincident with the highest temperatures measured in the at- mosphere. The properties of the broad polar cap are suggestive of polar vortices on other planets (13–16), but instead of anomalously cold temperatures, the southern polar temper- atures on Saturn are warm, sustained by nearly 15 years of continuous illumination. The exis- tence of a zonal jet at planetocentric latitude 69-S(17) is consistent with this interpretation. This explanation would predict a cold vortex at Saturn_s north pole, having undergone radia- tive cooling for nearly 15 years, a phenome- non that can be verified by Cassini_s thermal spectrometer CIRS in the near future. The appearance of a compact region of Fig. 3. Zonal temperature variations in the strato- maximum temperatures within 3- latitude of Fig. 2. Meridional variation of zonally averaged sphere (A) and troposphere (B). The temperatures the south pole is unexpected, having no long- properties of Saturn’s atmosphere. The top two at 46- and 57-S are offset to lower tempera- term analog in other planetary atmospheres panels show temperatures at 3 mbar (A) and tures at 1.5- and 3.5- latitude, respectively. Tro- (18). The enhanced temperatures could be 100 mbar (B). The extent of T2SDlimitsis pospheric temperatures cover nearly double the explained by local radiative heating by strato- shown in the 3-mbar plot, and the extent of longitude range of the stratospheric temper- spheric particulates entrained in near-polar T1 SD limits is shown in the 100-mbar plot. The atures as a result of the second mosaic taken at 17.65 mm. and polar regions, consistent with the increase two solid circles in (A) represent temperatures predicted by time-dependent radiative-convective of stratospheric particulates derived from anal- models (11, 12) at the equator and south pole ysis of cloud reflectivity (17), but we must still for the date of these observations. (C) The ver- shear; thus, the prograde wind speed gradient invoke dynamics to keep the stratospheric tical gradient of the zonal wind velocity, u,per increases with altitude, although not suffi- particles themselves entrained in such com- atmospheric scale height, ¯u/¯ln(p), at two pres- ciently to change the sign of the wind speed pact areas. Thus, dynamical as well as radi- sure levels, derived from the meridional gradient at 3 mbar. Between 20- and 50-S, the strato- ative forcing is required to explain the strong of temperature, assuming geostrophic balance. spheric wind shear is closer to zero than in temperature enhancement. In fact, heating the troposphere, consistent with a diminish- of the polar atmosphere by forced down- Between 20- and 50-S, there are no correla- ing wind with altitude. welling of relatively dry air is also consistent tions between tropospheric and stratospheric Most of the zonal bands are axisym- with the deeper tropospheric clouds sensed at temperatures or between temperatures and vis- metric, but zonal (longitudinal) variability is the pole (17). ible reflectivity. evident in the images (Fig. 1), and its pres- The organization of stratospheric and tropo- On the other hand, the instantaneous geo- ence can be detected by the standard devia- spheric temperatures over lower latitudes is strophic wind shear at 100 mbar, computed tion associated with zonal mean temperatures dominated by zonal bands of alternating warm from the thermal wind approximation for non- (Fig. 2). The highest amplitude zonal oscil- and cool temperatures, with temperature dif- zero latitudes, is correlated with prograde lations of the tropospheric temperature field ferences higher in the troposphere than the (positive, eastward) zonal cloud-tracked wind are near 32-S (Fig. 3). We analyzed the power stratosphere (Fig. 2). The area of lowest tropo- speeds (21–23) between 20- and 70-S. Near spectrum of these oscillations using a Lomb- spheric temperatures near the equator coin- 20-S, both the 3-mbar and 100-mbar wind Scargle periodogram, similar to earlier studies cides with a region of higher cloud reflectivity shears are negative, consistent with a Jupiter- of Jupiter (3, 4) and Saturn (20) which were and wind speed. The cold region has been like falloff of winds with altitude (24). How- characterized by irregular or incomplete longi- noted in earlier studies of Saturn_s tempera- ever from 5- to 7-S, the 3-mbar wind shear tudinal sampling. The power spectrum of this ture from Pioneer 11 (19) and Voyager 1 (20). has the opposite sign of the 100-mbar wind structure is dominated by wavenumbers 9 to

www.sciencemag.org SCIENCE VOL 307 4 FEBRUARY 2005 697 R ESEARCH A RTICLES 10. Similar zonal temperature variability is 3. G. S. Orton et al., Science 252, 537 (1991). 21. A. Sa´nchez-Lavega,R.Hueso,S.Pe´rez-Hoyos, J. F. Rojas, seen as far as 4- north and south of this lat- 4. G. S. Orton et al., Science 265, 625 (1994). R. G. French, Icarus 170, 519 (2004). 5. R. A. Hanel et al., Science 212, 192 (1981). 22. A. Sa´nchez-Lavega, J. F. Rojas, P. V. Sada, Icarus 147, itude at lower amplitude (Fig. 1). Zonal oscil- 6. R. A. Hanel et al., Science 215, 544 (1982). 405 (2000). lations of lower amplitude are also present 7. J. Pirraglia, B. J. Conrath, M. D. Allison, P. J. Gierasch, 23. A. Sa´nchez-Lavega,R.Hueso,S.Pe´rez-Hoyos, J. F. Rojas, in the troposphere and stratosphere at other Nature 292, 677 (1981). R. G. French, Nature 423, 623 (2003). 8. B. J. Conrath, P. J. Gierasch, E. Ustinov, Icarus 135, 24. P. J. Gierasch, J. A. Magalha˜es, B. J. Conrath, Icarus 67, latitudes (Fig. 3). The power spectra of these 501 (1998). 456 (1986). regions show no components rising above 9. F. M. Flasar et al., Science, published online 23 Decem- 25.J.A.Magalha˜es,A.L.Weir,P.J.Gierasch,B.J.Conrath, the 99% false-alarm probability level. ber 2004 (10.1126/science.1105806). S. S. Leroy, Icarus 88, 39 (1990) and references therein. 10. Details of the temperature retrieval process are 26. We thank the engineering staff of the W. M. Keck II Zonal temperature oscillations on Saturn described in the supporting material on Science Observatory, in particular LWS instrument special- were detected in Voyager 1 infrared maps (20), Online. ists R. Campbell and M. Kassis; B. Fisher of the Jet and a similar phenomenon has been seen on 11. B. Be´zard,D.Gautier,B.Conrath,Icarus 50, 274 Propulsion Laboratory (JPL) for his data reduction (1984). advice; A. Sanchez-Lavega and S. Perez Hoyos of the Jupiter (3, 4, 25). The oscillations on Jupiter 12. B. Be´zard, D. Gautier, Icarus 61, 296 (1985). Universitdad del Pais Vasco for their Hubble Space were shown to be thermal waves that are 13. See (14) for Earth’s Antarctic vortex. Telescope data and its reduction; and A. James Friedson moving slowly with respect to the rotation of 14. J. R. Schoeberl, D. R. Hartmann, Science 251, 46 (1991). for several helpful discussions about Saturn’s dy- 15. See (16) for Titan’s polar vortex. namics. Sponsored by a grant from NASA to JPL. The the interior of the planet and are uncorrelated 16. F. M. Flasar, Planet. Space Sci 46, 1125 (1998). tables required to look up temperatures as a func- with the zonal winds associated with Jupiter_s 17. A. Sa´nchez-Lavega,S.Pe´rez-Hoyos, J. R. Acarreta, tion of outgoing radiance and emission angle were cloud system (25); a similar conclusion was R. G. French, Icarus 160, 216 (2002). computed using a radiative-transfer code run efficient- reached regarding Saturn (20). Similar ground- 18. The closest analog is the sudden polar warming ly with the help of JPL’s Supercomputing Project. phenomenon in Earth’s atmosphere (see, for exam- based and Cassini CIRS observations will be ple,R.J.Reed,J.L.Wolfe,andH.Nishimoto.J. Atmos. Supporting Online Material needed to determine the phase speed of the Sci. 20, 2556, 1963), but it is a very short-term www.sciencemag.org/cgi/content/full/307/5710/696/ thermal waves. phenomenon, unlike Saturn’s warm polar region, as DC1 suggested by several years of lower-resolution ther- SOM Text mal imaging from the 3-m NASA Infrared Telescope Fig. S1 and S2 References and Notes Facility. References 1. F. C. Gillett, G. S. Orton, Astrophys. J. 195, L47 (1975). 19. A. P. Ingersoll, G. S. Orton, G. Munch, G. Neugebauer, 2. P. A. Yanamandra-Fisher, G. S. Orton, B. M. Fisher, S. C. Chase, Science 207, 434 (1980). 28 September 2004; accepted 10 December 2004 A. Sa´nchez-Lavega, Icarus 150, 189 (2001). 20. J. R. Achterberg, F. M. Flasar, Icarus 119, 350 (1996). 10.1126/science.1105730 REPORTS Sulfuric acid is a gas-phase particle pre- Rapid Formation of cursor that has been implicated for many years as a major atmospheric nucleating spe- Sulfuric Acid Particles at cies (5). Laboratory data on binary sulfuric acid nucleation rates with water, however, are insufficient by many orders of magnitude in Near-Atmospheric Conditions explaining atmospheric nucleation (6–8). Torsten Berndt,1* Olaf Bo¨ge,1 Frank Stratmann,1 Ternary nucleation, involving the additional Jost Heintzenberg,1 Markku Kulmala2 ubiquitous species ammonia, has been sug- gested from theoretical considerations as an We investigated the formation of new particles in a laboratory study, starting explanation for observed particle nucleation from H SO produced in situ through the reaction of OH radicals with in the atmosphere (9, 10). Further possible 2 4 routes for atmospheric particle formation are SO2. Newly formed particles were observed for H2SO4 concentrations above 7 106 per cubic centimeter. At 293 kelvin, a rough estimate yielded a nucleation processes involving oxide nucleation rate of 0.3 to 0.4 particles per cubic centimeter per second for for coastal areas (11, 12), ion-induced È 7 Q nucleation (13), and nucleation assisted by 10 particles per cubic centimeter of H2SO4 (particle size 3 nanometers). These findings are in agreement with observations from the atmosphere. The products from the atmospheric oxidation of results demonstrate that under laboratory conditions similar to the atmo- aromatics (14). sphere, particle formation occurs at atmospheric H SO concentration levels. We report results from laboratory experi- 2 4 ments conducted in an atmospheric pressure Mass balances of the atmospheric aerosol several reasons, the source processes control- flow-tube that was irradiated with ultraviolet have been used for more than 30 years (1)to ling the number of secondary particles in the (UV) lamps when required (15). In our first

identify potential human influence on cli- atmosphere are much more complex and dif- experiments, we took H2SO4 from a liquid mate. This is appropriate for mass-dominated ficult to understand than those of the primary reservoir by passing a part of the dry carrier

sources of primary aerosol particles, such as particles: (i) a host of gas-phase particle pre- gas through a H2SO4 saturator (97.5%, Fluka), Earth_s crust or the sea surface, and their cursors with their respective formation pro- and mixed this gas stream with humidified influence on processes controlled by par- cesses is involved; (ii) particle nucleation carrier gas at the entrance of the flow tube. ticulate mass, such as iron fertilization of from the gas phase may possibly comprise 1Leibniz-Institut fu¨r Tropospha¨renforschung e.V., southern oceans (2). On the other hand, several species concurrently, including ions; 2 Permoserstraße 15, 04318 Leipzig, Germany. Uni- crucial processes such as cloud formation and (iii) there are no experimental tech- versity of Helsinki, Post Office Box 64, FIN-00014, (3) and possibly even certain health effects niques available to size, quantify, or chem- Finland. (4) are controlled by the number of aerosol ically specify the newly formed particles ab *To whom correspondence should be addressed. particles, including secondary particles. For initio. E-mail: [email protected]

698 4 FEBRUARY 2005 VOL 307 SCIENCE www.sciencemag.org R EPORTS

Particle formation took place for H2SO4 con- and 3. The corresponding rate coefficients is supported by experiments using ozonolysis centrations above 1010 cmj3, which is in line were taken from literature (20–23). of olefins as a Bdark[ OH radical source (26). with observations from other researchers using In Fig. 2, experimentally observed particle Formation of new particles was detected for È 7 j3 0 H2SO4 directly from a liquid reservoir (6–8). numbers for calculated H2SO4 concentrations 2 10 cm of H2SO4 at r.h. 28%, nearly Further experiments applied in situ gas- are plotted with different hydrocarbons (cyclo- independently of the olefin used. In these phase formation of H2SO4 in the flow tube hexane, n-heptane, mesitylene, and furan) for experiments, the reaction pattern describing E 0 (Fig. 1). The chosen pathways leading to OH radical titration relative humidity (r.h.) H2SO4 formation was more complex and not ^ H2SO4 are equal to those in the atmosphere 28% . The stated H2SO4 concentration is all kinetic coefficients were available, making 1 (16). O3 was photolyzed, forming O( D) calculated for the end of the irradiated zone. rough estimates necessary. Considering the pos- atoms, and the subsequent reaction with A concentration profile inside the flow tube is sible uncertainties in these experiments, e.g., a water vapor produced OH radicals. The OH given in fig. S1. In contrast to the experi- factor of 2 to 4 with respect to the calculated radicals attacked SO2, and after the reaction ments using H2SO4 from a liquid reservoir, in H2SO4, the agreement of the results from ozon- 10 j3 with O2, the adduct HOSO2 yielded SO3 and which particle formation started at 10 cm olysis (26) with those from photolysis is good. HO2. In the reaction of SO3 with water va- of H2SO4, here formation of new particles For a fixed H2SO4 concentration, the num- por, two molecules of H2O or one dimer of had already begun at H2SO4 concentrations of ber of new particles increased for increasing 6 j3 H2O per molecule of SO3 are needed (17, 18), 7 10 cm . According to a rough estimate, r.h. (Fig. 3). The increase was much more 0 which finally led to H2SO4. A hydrocarbon the uncertainty of these calculated H2SO4 con- pronounced for the lowest r.h. values. At r.h. È 7 j3 (at first furan) was added for OH radical centrations is about a factor of 2. The slopes of 11%, 4 10 cm of H2SO4 were needed titration. Formation of new particles was the particle number versus H2SO4 concentra- to produce detectable numbers of particles. For visible only in the presence of all reactants tion curves (here between 3 and 5 depending higher r.h. values (28% and 49.5%), the dif- needed for H2SO4 formation, including UV on r.h.) give the number of the H2SO4 mole- ferences in the number of formed particles radiation for OH radical production. The lack cules in the critical cluster according to the were small, close to the uncertainty of the of particles in the absence of SO2 indicates nucleation theorem (24). However, this num- measurement. Previous studies with H2SO4 that the reaction of OH radicals with furan ber might be overestimated. This is because (i) from a liquid reservoir revealed similar r.h. does not produce new particles itself. This particle diameters measured at the end of dependencies, especially for low r.h. values fact was confirmed for all hydrocarbons ap- the flow tube are È3 nm and increase with (6, 8). plied. The lack of particles in the absence increasing H2SO4 concentration; and (ii) for Atmospheric data from simultaneous mea- È of O3 indicates that no further photolytic pro- particle diameters 3 nm, the particle detec- surements of newly formed particles and cesses are forming particles. tion efficiency increases with increasing par- H2SO4 at ground level are rather rare. How- To determine the H2SO4 concentration in ticle size. The combination of these two facts ever, existing data sets show consistently that È 7 j3 the flow tube, we used a simple kinetic model. yields increasing detection efficiencies with in- 10 cm of H2SO4 are necessary in the creasing H SO concentrations, i.e., artificially atmosphere for particle formation (27–29). þ ð Þ Y ð Þ 2 4 OH HC furan products 1 larger slopes in Figs. 2 and 3. This is exactly the order of magnitude of The data points obtained for the different OH þ SO2 Y I: Y H2SO4 ð2Þ hydrocarbons show very good agreement, in- dicating that the different reaction products from pathway 1 do not participate in the nu- H2SO4 Y wall ð3Þ cleation process substantially or influence Initial concentrations of the hydrocarbons the nucleation process in a nearly analo- (HC; cyclo-hexane, n-heptane, mesitylene, gous manner. Reaction products arising from and furan) and SO2 were chosen so that more cyclo-hexane are mainly cyclo-hexanone and than 99% of the OH radicals reacted via cyclo-hexanol (25), with final concentrations pathway 1. That allowed the determination of on the order of magnitude of 1011 cmj3 or the OH radical concentration in the irradiated less. For n-heptane, mesitylene, and furan, zone of the flow tube from the measured information in the literature concerning pro- amount of consumed HC (19). With this duct formation is sparse. knowledge, H2SO4 concentrations could be The finding that particle formation takes È 7 j3 easily calculated by considering pathways 2 place for H2SO4 concentrations of 10 cm

Fig. 1. Measured particles (particle size Q 3nm) from a typical experiment for different trace gas additions, at 293 K, r.h. 0 11%. The graph has an offset of 0.01 particles cmj3. Initial concentra- 0 tions were [O3] 4.6 Fig. 2. Experimentally observed particle 11 j3 0 10 cm , [SO2] 8.1 numbers for different hydrocarbons (cyclo- 1010 cmj3, and [furan] 0 hexane, n-heptane, mesitylene, and furan) for 1.1 1012 cmj3. OH radical titration, at r.h. 0 28%. Initial 0 concentrations of the reactants were [O3] 11 j3 0 (4.4 to 9.8) 10 cm , [SO2] (5.9 to 81) 109 cmj3, and [HC] 0 (1.1 to 11) 1012 cmj3. The amount of converted HC was in the range (7.9 to 31) 1010 cmj3, and the OH radical concentrations were (8.4 to 18) 106 cmj3.

www.sciencemag.org SCIENCE VOL 307 4 FEBRUARY 2005 699 R EPORTS

7 j3 H2SO4 where we observed particle formation zone is below 10 cm (0.5 pptv). Initial as References and Notes using in situ–produced H SO . Derived parti- well as averaged NH concentrations in the 1. C. Wilson, Ed., Inadvertent Climate Modification, 2 4 3 Report of the Study of Man’s Impact on Climate cle nucleation rates in the continental atmo- nucleation zone are distinctly lower than at- (MIT Press, Cambridge, MA 1971). sphere at ground level are in the range 0.5 to mospheric NH3 mixing ratios measured over 2. A. J. Watson, D. C. E. Bakker, A. J. Ridgwell, P. W. 10 particles cmj3 sj1 (27–30). continents of 100 to 10,000 pptv (34). Boyd, C. S. Law, Nature 407, 730 (2000). 3. H. R. Pruppacher, J. D. Klett, Microphysics of Clouds and From the experiments with in situ H2SO4 In the experiments with in situ–produced Precipitation (Reidel, Dordrecht, Netherlands, 1978). formation, we estimated a nucleation rate of H2SO4 (and in the presence of hydrocarbons), 4.A.Peters,H.-E.Wichmann,T.Tuch,J.Heinrich,J.Heyder, 0.3 to 0.4 particles cmj3 sj1 for 107 cmj3 of particle formation occurs for È107 cmj3 of Am.J.Respir.Crit.CareMed.155, 1376 (1997). 0 5. G. J. Doyle, J. Chem. Phys. 35, 795 (1961). H2SO4 and r.h. 28% to 49.5% (Fig. 3), H2SO4 with a nucleation rate of 0.3 to 0.4 6. B. E. Wyslouzil, J. H. Seinfeld, R. C. Flagan, K. Okuyama, j j using calculated concentration profiles (fig. particles cm 3 s 1 (both parameters compa- J. Phys. Chem. 94, 6842 (1991). S1) for defining the nucleation zone (15). This rable to those from the atmosphere), but at 7. Y. Viisanen, M. Kulmala, A. Laaksonen, J. Chem. Phys. finding is in line with the lower limit of the distinctly lower NH concentrations than in 107, 920 (1997). 3 8. S. M. Ball, D. R. Hanson, F. L. Eisele, P. H. McMurry, nucleation rates observed in the atmosphere. the atmosphere. For this reason, a substantial J. Geophys. Res. 104, 23709 (1999). 9. D. J. Coffman, D. A. Hegg, J. Geophys. Res. 100, 7147 Ternary nucleation using H2SO4,H2O, role of NH3 in the nucleation process becomes and NH (9, 10) is often assumed in descrip- questionable. Furthermore, the same carrier gas (1995). 3 10. P. Korhonen et al., J. Geophys. Res. 104, 26349 (1999). tions of new particle formation in the atmo- was used in both the experiments with in situ– 11. C. D. O’Dowd et al., Nature 417, 632 (2002). 12. C. E. Kolb, Nature 417, 597 (2002). sphere. Experimental evidence has been found produced H2SO4 and those with H2SO4 from for the formation of clusters consisting of a liquid reservoir. This makes NH influence 13. S. H. Lee et al., Science 301, 1886 (2003). 3 14. R. Zhang et al., Science 304, 1487 (2004). H2SO4 and NH3 for temperatures up to 285 K, on the barrier H2SO4 concentrations needed 15. Materials and methods are available as supporting given concentrations of È109 cmj3 for both for nucleation unlikely, because for both material on Science Online. species (31). In our experiment, no NH was experiments, the carrier gas should contain 16. B. J. Finlayson-Pitts, J. N. Pitts Jr., Chemistry of the Upper 3 and Lower Atmosphere: Theory, Experiments, and added. The measured NH3 concentrations in the same unmeasurable amount of NH3. Applications (Academic Press, San Diego, CA, 2000). the carrier gas were below the detection limit We compared our experimental nuclea- 17. E. R. Lovejoy, D. R. Hanson, L. G. Huey, J. Phys. Chem. (3.5 109 cmj3) of the AiRRmonia system tion rates with theoretical ones (35, 36). The 100, 19911 (1996). 18. J. T. Jayne et al., J. Phys. Chem. 101, 10000 (1997). used (32). We modeled the NH3 concentra- H2SO4 concentration needed for substantial 19. Here a nearly constant OH radical concentration in j tion profile in the tube by taking this value as binary nucleation is È1010 cm 3, i.e., far too the irradiated reaction zone is assumed. This is valid for a constant ratio of OH formation rate to OH con- the initial NH3 concentration at the entrance high compared to the experimental values. sumption rate to be proportional to [O3]/[HC]. Limiting of the flow tube and assuming diffusion- Ternary NH3-influenced nucleation does not the conversion of O3 and HC to 10 to 15% makes controlled wall loss for NH3 (33). explain the observed particle numbers, if NH3 the assumption of constant OH radical levels viable. mixing ratios below 0.5 pptv are assumed (as 20. R. Atkinson, Chem. Rev. 86, 69 (1986). Y ð Þ modeled above). Furthermore, the nucleation 21. F. Kramp, S. E. Paulson, J. Phys. Chem. 102 A, 2685 NH3 wall 4 (1998). rate seems to behave like H2SO4 concentra- 22. R. Zellner, Ber. Bunsenges. Phys. Chem. 82, 1172 (1978). 23. D. R. Hanson, F. L. Eisele, J. Phys. Chem. 104 A, 1715 This modeling results in a maximum NH3 tion to a power of smaller than 3 to 5, indi- concentration of È2 108 cmj3 E8 parts per cating that the nucleation mechanism depends (2000). ^ 24. D. Kashchiev, J. Chem. Phys. 76, 5098 (1982). trillion by volume (pptv) at the beginning of clearly on H2SO4 concentration, and suggest- 25. D. M. Rowley, P. D. Lightfoot, R. Lesclaux, T. J. Wallington, the nucleation zone, dropping substantially ing a very close to trimer or dimer (kinetically J. Chem. Soc., Faraday Trans. 1 87, 3221 (1991). with increasing residence time. The modeled controlled) nucleation mechanism (37). 26. T. Berndt, O. Bo¨ge, F. Stratmann, Atmos. Environ. 38, 2145 (2004). average NH3 concentration in the nucleation No definite mechanistic explanation for 27. R. J. Weber et al., Geophys. Res. Lett. 26, 307 (1999). the different barrier H2SO4 concentrations of 28. W. Birmili, A. Wiedensohler, C. Plass-Du¨lmer, H. È107 cmj3 and È1010 cmj3 can be given. Berresheim, Geophys. Res. Lett. 27, 2205 (2000). 29. W. Birmili et al., Atmos. Chem. Phys. 3, 361 (2003). Currently available binary nucleation theories 30. M. Kulmala et al., J. Aerosol Sci. 35, 143 (2004). are not able to describe these findings and 31. D. R. Hanson, F. L. Eisele, J. Geophys. Res. 107, may not consider the correct nucleation mech- 10.1029/2001JD001100 (2002). anism. Ion-induced nucleation also cannot 32. J. W. Erisman et al., Atmos. Environ. 35, 1913 (2001). 33. The reactor wall is assumed to be coated with a explain our experimental results, because no H2O/(H2SO4) layer. Uptake coefficients for NH3 on ions were present in the experiments. Argu- neutral or acidic solutions are in the range 0.05 to ments against NH -influenced ternary nu- 1.0 (39), making the assumption of diffusion- 3 controlled wall loss reasonable. The diffusion co- cleation include (i) different barrier H2SO4 efficient was taken from Lechner (40). concentrations for nucleation of È107 cmj3 34. J. H. Seinfeld, S. N. Pandis, Atmospheric Chemistry and È1010 cmj3 despite approximately the and Physics (Wiley, New York, 1998). 35. H. Vehkamaki et al., J. Geophys. Res. 107, 4622, same anticipated NH3 concentration in the car- doi:10.1029/2002JD002184 (2002). rier gas, and (ii) a too-low NH3 concentration 36. I. Napari, M. Noppel, H. Vehkamaki, M. Kulmala, in the nucleation zone. Tentatively, it can be J. Chem. Phys. 166, 4221 (2002). 37. A. A. Lushnikov, M. Kulmala, Phys. Rev. E58, 3157 (1998). estimated that the difference in the barrier 38. M. Kalberer et al., Science 303, 1659 (2004). H2SO4 concentration is due to effects con- 39. E. Swartz et al., J. Phys. Chem. 103 A, 8824 (1999). 40. M. D. Lechner, Taschenbuch fu¨r Chemiker und Physiker nected with freshly produced H2SO4 and/or the hydrocarbon_s influences on the nucleation (Springer-Verlag, Berlin, 1992). 41. We thank A. Gru¨ner and G. Spindler for NH3 mea- and/or the subsequent growth processes. If surements, M. Schu¨tze for helpful discussions, and hydrocarbons bear a meaning for the observed K. Pielok and H. Macholeth for technical assistance. particles, the oxygenated reaction products Supporting Online Material from pathway 1 (carbonylic substances, etc.) www.sciencemag.org/cgi/content/full/307/5710/698/ DC1 are probably more suitable candidates than Fig. 3. The number of newly formed particles Materials and Methods for r.h. 0 11%, 19%, 28%, and 49.5%, given their precursor molecules (14, 35). More work Fig. S1 is needed to clarify the mechanism leading to furan for OH radical titration (in situ H2SO4 13 August 2004; accepted 27 December 2004 formation in the presence of hydrocarbons). the particles observed. 10.1126/science.1104054

700 4 FEBRUARY 2005 VOL 307 SCIENCE www.sciencemag.org R EPORTS number of possible Laves phases derives from the fact that there are two forms of these Dislocations in Complex Materials three-layer stacks that we designate as t t¶ - 1 2 3 and , which are rotated 180 in the layer Matthew F. Chisholm, Sharvan Kumar, * Peter Hazzledine plane to each other. In C14, these two ar- rangements of the three-layer stacks alter- Deformation of metals and alloys by dislocations gliding between well- nate (Fig. 1A) and the stacking sequence of separated slip planes is a well-understood process, but most crystal structures single basal planes (s) and triple basal planes do not possess such simple geometric arrangements. Examples are the Laves (t)isstst¶I . All other Laves phases have phases, the most common class of intermetallic compounds and exist with longer stacking sequences than C14, and thus, ordered cubic, hexagonal, and rhombohedral structures. These compounds are during a transformation, some t layers must usually brittle at low temperatures, and transformation from one structure to be changed to t¶ and vice versa; this may be another is slow. On the basis of geometric and energetic considerations, a accomplished by passage of the transform- dislocation-based mechanism consisting of two shears in different directions ing Shockley dislocations, much like in the on adjacent atomic planes has been used to explain both deformation and fcc structure. phase transformations in this class of materials. We report direct observa- An ideal technique for viewing the atomic tions made by Z-contrast atomic resolution microscopy of stacking faults and structures in Laves phases is Z-contrast imaging dislocation cores in the Laves phase Cr2Hf. These results show that this com- in a high-resolution scanning transmission elec- plex dislocation scheme does indeed operate in this material. Knowledge gained tron microscope (STEM) (12). With its sub- of the dislocation core structure will enable improved understanding of de- angstrom resolution (13), the positions of atom formation mechanisms and phase transformation kinetics in this and other columns are revealed, and, because the atomic complex structures. number EZ^ values for Cr E24^ and Hf E72^ are very different, the light and heavy atoms may Plastic deformation in a crystal occurs by the nisms based on dislocation motion have been be distinguished. Figure 1B is a micrograph of G 9 processes of slip and twinning. These pro- proposed (2, 3, 5, 9). C14 Cr2Hf viewed along 11-20 in the same cesses are accomplished by the motion of dis- On an atomic level, however, the shear- orientation as Fig. 1A. locations whose character is closely related to ing mechanisms in the Laves phases can- It is known from conventional TEM ob- the structure of the crystal. When these dislo- not be identical to those in simple metals servations that the slip planes for the cations produce displacements that are less because Laves phases are ordered struc- Shockley dislocations in the C14 structure than a unit lattice translation vector in the tures with four different atomic planes par- are the basal planes (4, 6), although atomic crystal, they are called partial dislocations allel to the slip plane (10). A ball-and-stick details remain unresolved. Slip, twinning, and they bound stacking faults. The motion of model of the hexagonal C14 structure (the and shear transformations are believed, on a partial dislocation can also produce certain Laves phase with the shortest period along geometric and energetic grounds, to occur types of phase transformations. The partial the basal plane normal) when viewed along inside the three-layer stack (rather than dislocation associated with the sliding of a close-packed direction and with basal planes between the single Cr layer and the three- close-packed planes of atoms over each other horizontal is illustrated (Fig. 1A). The al- layer stack) by a mechanism called syn- during slip, twinning, or shear transforma- loy Cr2Hf is used as the example. The chroshear (3, 4, 14) that consists of two tions in face-centered cubic (fcc) metals is the structure, like all Laves phases, consists of shears in different directions on adjacent Shockley partial (1). In slip, a pair of partial alternating layers of single small (Cr) atom atomic planes. The dislocation description dislocations bounding a stacking fault moves layers Ein the form of a kagome (basket of synchroshear requires the passage of two on the slip plane in response to an applied weave) net (11)^ and three-layer stacks (Hf- Shockley dislocations through the three-layer stress to produce plastic deformation. In Cr-Hf) that contain two low-density planes stack, one gliding between the lower atom twinning, a Shockley dislocation sweeps of large (Hf) atoms separated by a low- level and the center with one Burgers vector every slip plane, whereas when a Shockley density plane of small (Cr) atoms. The great and the other gliding between the center and dislocation sweeps alternate slip planes it converts an fcc structure into a hexagonal close-packed (hcp) structure (1). Observa- tions on Laves phases (2) show that analo- gous mechanisms could operate in these more complex structures (3–7). Laves phases, compounds with the AB2 stoichiometry where the large A atoms and the small B atoms have an ideal radius ratio of 1.225, are the most commonly occurring intermetallic compounds. Cubic, hexagonal, and rhombo- hedral structures have been imaged with the use of high-resolution electron microscopy (2–4, 8), and phase transformation mecha-

1Condensed Matter Sciences Division, Oak Ridge National Laboratory, Oak Ridge, TN 37831, USA. 2Division of Engineering, Brown University, Provi- G 9 3 Fig. 1. (A) A schematic of the 11-20 projection of the C14 hexagonal structure showing the three- dence, RI 02912, USA. UES, Incorporated, Dayton, OH layer stack, t,andthe180--rotated three-layer stack, t¶, separated by the single layer, s. The large 45432, USA. atoms (Hf) are in yellow and the small atoms (Cr) are in red. (B) A Z-contrast image of the G11-209

*To whom correspondence should be addressed. projection of the C14 variant of the Cr2Hf Laves phase. The bright features (yellow) are Hf columns, E-mail: [email protected] and the less-intense features (red) are Cr columns.

www.sciencemag.org SCIENCE VOL 307 4 FEBRUARY 2005 701 R EPORTS the upper level with a different Burgers Fig. 2. A schematic illus- vector. Figure 2 illustrates the sliding of tration of the motion of a two rigid blocks, whose faces terminate at synchro-Shockley partial dislocation. (A) The initial the upper and lower Hf layers in the three- arrangement of the big layer stack, past each other in response to an (yellow) and small (red) applied stress, with an intermediate Cr layer atoms in the three-layer between the two blocks. If it is assumed that stack, shown with the the bottom block is stationary, then in re- same orientation as in sponse to an applied shear stress the Hf Fig. 1. The kagome layers above and below the atom in the top block moves in the plane three-layer stack are rep- of the paper in the direction shown by the resented by the rigid bold black arrow to the position of the Cr blocks, with the bottom atom labeled X. For this to happen, however, block considered station- the Cr atom in the layer between the two ary and the top block rigid blocks needs to get out of the way, and responding to a shear stress by moving to the it does so by moving out of the plane of the left. This representation is paper in the direction illustrated by the blue the motion of a synchro- arrow to position X ¶, which was previously Shockley partial dis- occupied by a Hf atom. These two co- location with pure edge ordinated movements can be described by character. (B)Theatomic the motion of two Shockley partial disloca- arrangement after syn- chroshear. (C)Thesyn- tions that for energetic reasons are closely chroshear process is coupled. For this reason, in the literature, the illustrated where all the pair of Shockley dislocations has been Hf atoms in the top treated as a single synchro-Shockley partial layer of the three-layer dislocation with its core spread on two stack move in the plane of the paper to the left by a distance b as shown by the bold black arrow, while - adjacent planes (9). The effect of this syn- the Cr atoms move out of the plane of the paper in the middle Cr layer (at 60 to the –X direction) as shown by the blue arrow, producing the final configuration shown in (B). chronous translation of the two layers is to convert a t-type three-layer stack into a t¶- type three-layer stack, thereby producing a Fig. 3. A Z-contrast image of the G11-209 projection of a synchro- stacking fault. In Fig. 2, the synchro- Shockley dislocation bounding a Shockley partial dislocation is an edge stacking fault in the C14 variant dislocation. of the Cr2Hf Laves phase. The The Z-contrast image in Fig. 3 shows a fault comes in from the right (indicated by the arrows) and stacking fault in Cr2Hf that comes in from the right about halfway up the image (indicated terminates at the dislocation core in the center of the image. by the arrow) and terminates in the center of The synchro-Shockley partial dis- the micrograph at the core of a synchro- location in this instance has a 30- Shockley dislocation. The construction of a character. The Burgers circuit Burgers circuit using the Hf atoms in the made from Hf atoms failed to triple layer shows closure failure. In the core close. The schematic below the of the dislocation, a t layer (on the left) image, obtained directly from the t¶ image, shows a proposed core changes to a layer (on the right). Relative structure. The indicated Cr col- to the lower Hf atoms, the upper Hf atoms umns in the small box in the sche- may move either to the right by b/2 (where b matic are included on the basis is the Burgers vector), signifying a 30- char- of the relatively large separation acter for the dislocation, or to the left by b of the adjacent Hf columns. (as in Fig. 2), which would imply a pure edge dislocation (90- character). The closure failure of the Burgers circuit in Fig. 3 is 0.15 nm 0 b/2, implying that this dislocation has a 30- character. As may be seen in the figure, the core of the synchro-Shockley dis- location is compact, on the order of 0.5 nm in width and about 0.1 nm high. 4is....st¶stst¶st¶st¶st¶st¶stst¶.... instead of the (4) for Laves phases^ actually occurs in the ...st¶stst¶stst¶stst¶stst¶... A Laves phase transformation is illus- unsheared . Cr2Hf Laves phase. Because all Laves phases trated (Fig. 4) where a strip of the cubic The concept of synchroshear described are constructed from the same building C15 Laves phase has formed in the C14 above was originally introduced by Kronberg blocks, there is good reason to believe that structure by repeated synchroshear. The C15 (14) to explain the deformation behavior of synchroshear occurs in the others as well.

structure results by synchroshearing the C14 a-Al2O3, although recent results (15–17) The motion of a single type of defect, a structure on alternate three-layer stacks; this have confirmed that that is not the case. The synchro-Shockley partial dislocation, is is analogous to ordinary Shockley partial Z contrast images of faults and dislocations thought to be responsible for shear phase dislocations transforming a hcp structure to a presented here, however, demonstrate unam- transformations as well as three mechanisms fcc structure, by gliding on alternate close- biguously that synchroshear Eoriginally pro- of deformation: slip, twinning, and stress- packed planes. The stacking sequence in Fig. posed by Allen, Delavignette, and Amelinckx induced transformations. In this case, the

702 4 FEBRUARY 2005 VOL 307 SCIENCE www.sciencemag.org R EPORTS Fig. 4. A Z-contrast image of the 4. C. W. Allen, P. Delavignette, S. Amelinckx, Phys. Status G11-209 projection of a faulted region Solidi A 9, 237 (1972). 5. Y. Liu, J. D. Livingston, S. M. Allen, Metall. Mater. of the C14 variant of the Cr2Hf Laves phase. This region (between the dou- Trans. A 23, 3303 (1992). 6. K. S. Kumar, D. B. Miracle, Intermetallics 2, 257 (1994). blearrowontheleft)hastheC15 7. Y. Liu, J. D. Livingston, S. M. Allen, Met. Mat. Tran. A cubic structure and could be formed 26, 1441 (1995). by stacking faults in the C14 struc- 8. Y. Kitano, Y. Komura, H. Kajiwara, E. Watanabe, Acta ture (indicated by the two short Crystallogr. A 36, 16 (1980). arrows on the right) that result from 9. P. M. Hazzledine, P. Pirouz, Scripta Metall. Mater. 28, the passage of two synchro-Shockley 1277 (1993). dislocations. 10. C. S. Barrett, T. B. Massalski, Structure of Metals: Crystallographic Methods, Principles, and Data, vol. 35 of International Series on Materials Science and Tech- nology (Pergamon, Oxford, ed. 3,1987). 11. K. Husimi, Prog. Theor. Phys. 5, 177 (1950). 12. P. D. Nellist, S. J. Pennycook, Phys. Rev. Lett. 81, 4156 (1998). 13. P. D. Nellist et al., Science 305, 1741 (2004). 14. M. L. Kronberg, Acta Metall. 5, 507 (1957). 15. J. B. Bilde-Sørensen et al., Acta Mater. 44, 2145 (1996). 16. P. Pirouz et al., Acta Mater. 44, 2153 (1996). 17. T. Geipel et al., Acta Mater. 44, 2165 (1996). 18. This research was sponsored by the Office of Basic Energy Sciences, U.S. Department of Energy at Oak kinetics of all four of these processes is con- In addition, these concepts can be extended to Ridge National Laboratory, under contract no. DE- trolled by the mobility of synchro-Shockley other crystal structures where the slip planes AC05-00OR22725. S.K. acknowledges the support from the NSF-sponsored Materials Research Science dislocations. Much understanding could be have more than one spacing. and Engineering Center on Micro- and Nano-Mechanics gained from atomistic simulations of the of Materials at Brown University (contract no. DMR- References and Notes motion of this type of dislocation. Under- 9632524), and P.H. acknowledges support from U.S. 1. J. P. Hirth, J. Lothe, Theory of Dislocations (McGraw- Air Force Research Laboratory contract no. F33615- standing the structure of the dislocation core Hill Series in Materials Science and Engineering, 01-5214 with UES, Incorporated. and its influence on the ability of Laves McGraw-Hill, New York, 1968). 2. F. Laves, H. Witte, Metallwirtsch. Metallwiss. Metalltech. phases to deform is central to designing 15, 840 (1936). 4 October 2004; accepted 22 December 2004 alloys with optimized mechanical properties. 3. J. D. Livingston, Phys. Status Solidi A 131, 415 (1992). 10.1126/science.1105962

away from the end of a 1D structure can be End States in One-Dimensional answered directly. So far, definitive spectroscopic evidence Atom Chains for the existence of end states in 1D struc- tures has been lacking. Research has instead J. N. Crain* and D. T. Pierce focused on the quantum mechanics of elec- trons confined to reduced dimensions within End states—the zero-dimensional analogs of the two-dimensional states that the surface layer. Electrons are partially re- occur at a crystal surface—were observed at the ends of one-dimensional flected by step edges on metal surfaces and atom chains that were self-assembled by depositing gold on the vicinal Si(553) can exhibit refraction at the interface between surface. Scanning tunneling measurements of the differential two media (4–7). Electrons also exhibit quan- conductance along the chains revealed quantized states in isolated segments tum confinement when trapped within atomic with differentiated states forming over end atoms. A comparison to a tight- Bcorrals[ assembled by STM (8). At the 1D binding model demonstrated how the formation of electronic end states trans- limit, finite atomic chains of Au on NiAl(110) forms the density of states and the energy levels within the chains. (9, 10) and Cu on Cu(111) (11) constructed by STM exhibit quantized electronic states, The break in translational symmetry at a crys- duced dimensions at the edge or end of a and the observation of spectroscopic enhance- tal surface creates surface electronic proper- nanostructure. Similar to a two-dimensional ment at or near the ends of these chains ties that differ from those in the bulk crystal (2D) surface state formed at the surface of a suggested the possibility of the formation of and localize at the surface layer (1, 2). The bulk sample, an edge or step in a 2D struc- end states. However, the weakness of these formation of surface states and resonances is ture breaks the 2D symmetry and can form a effects in the systems studied, compounded a general property of solid surfaces, and the 1D edge state (3). Likewise, a finite 1D chain with the possibility of experimental artifacts delicate interplay between the surface elec- of atoms should exhibit zero-dimensional end (10, 11), precluded unequivocal assignment. tronic structure and the atomic positions often states at its termini. An end state requires two In the Si(553)-Au atom chains described be- leads to complex surface reconstructions in criteria: (i) The wave function of the state low, end states are unequivocally manifested which the atoms in the top layer rearrange to must be localized to the end atoms, and (ii) it through a marked transfer of the DOS from minimize the surface energy. must decay exponentially into the chain (2). the empty to the filled states above the end In analogy to the surface of a bulk solid, Scanning tunneling microscopy (STM) and atoms. Furthermore, we found that quantized we expect to observe similar physics in re- scanning tunneling spectroscopy (STS) enable states within finite chains can no longer be the spatial mapping of differentiated electron- described by a particle-in-a-box model, which ic structure within a nanostructure. Thus, the has been successful in the previous studies Electron Physics Group, National Institute of Stan- dards and Technology, Gaithersburg, MD 20899–8412, local density of states (DOS) can be mapped in (9–11), but rather require a model that in- USA. real space, providing direct access to electronic cludes end states. *To whom correspondence should be addressed. edge or end effects. Furthermore, questions To fabricate 1D chains, we used the self- E-mail: [email protected] of the spatial variation of the wave function assembly of chain reconstructions on stepped

www.sciencemag.org SCIENCE VOL 307 4 FEBRUARY 2005 703 R EPORTS Si templates driven by the deposition of gold (Fig. 1, B, D, and F). This apparent change states, we compared spectra from the center at elevated temperatures (12). Gold chain re- in length is caused by a contrast reversal over and edge of the dimer (Fig. 2D, red). At the constructions on stepped Si have been studied the end atoms, as is apparent from an en- center, the state at –0.75 V is clearly seen but extensively with angle-resolved photoemis- largement of a nine-atom chain (Fig. 1, C and the state at –0.45 V appears only as a shoulder. sion (12–16). Because photoemission gives D) and from topography line profiles along Over the edge of the dimer, although the in- spatially averaged electronic properties, to date the chain (Fig. 1, E and F). At positive bias tensity is low, the –0.75-V peak is absent and the role of individual defects has remained the end atoms are hardly visible and the atoms allows clear resolution of the –0.45-V state. largely unexplored (17, 18). Here, we focus on second from the end are enhanced, whereas In contrast, the three-atom chain exhibits Si(553)-Au, which exhibits several metallic at negative bias the end atoms are enhanced. a new peak that is nearly 1 eV higher in en- bands with 1D dispersion (19) and has few Such a polarity contrast in STM indicates ergy and is confined to the central atom. This defects. In this case, the defects appear as an underlying difference in the DOS for the apparent departure for the three-atom chain dark voids in STM images (Fig. 1) that leave empty and filled states near the ends of the is attributable to end states, which are char- isolated segments with chain lengths that vary chains. On the end atoms, the DOS is trans- acteristic of chains three atoms or more in

depending on the defect density. All of the ferred from the empty to the filled states. length. The empty state 0.5 eV above EF lies measurements were made at room tempera- The differential conductivity as measured in the gap of the Si substrate (Fig. 2B), but ture, higher than the temperatures at which by STS provides a direct measure of the lo- states below –0.3 eV are degenerate with the charge density wave transitions have been ob- cal DOS at the tip position and thus allows a Si bulk bands and are thus technically Bend served in similar 1D chains on Si (20, 21). detailed study of the electronic variation near resonances,[ analogous to the distinction be- Such transitions transform the electronic struc- the ends of a chain. Figure 2 shows an exam- tween states and resonances at surfaces of ture from metallic to insulating, so a room- ple for a seven-atom chain with spectra taken bulk solids. temperature study is desirable to access the beyond the end of the chain, over an end To elucidate these end effects, we per- chains in their metallic phase. atom, and on an interior atom. Beyond the end formed spectroscopy in a dense line of points The formation of end states in finite chain of the chain, there is a clear gap with little along three-, four-, and five-atom chains (Fig. 3,

segments is already apparent from compar- intensity near the Fermi energy (EF) (Fig. 2B, D, F, and H). These conductance images are ing STM images of the same sample region green). The boundaries of the gap are consist- formed from individual conductance spectra taken at opposite polarity (Fig. 1). At a sam- ent with the measured valence-band maximum like those in Fig. 2, B and D, taken at each ple bias of þ0.5 V (Fig. 1, A, C, and E), the for Si(553)-Au, –0.3 eV, from photoemission point along the atom chain. Each spectrum is chains appear shorter than at a bias of –1 V (12) and the 1.1-eV band gap of Si. Over the divided by a second-order polynomial, which end atom, the differential conductance ex- approximately removes the transmission func- hibits a new peak at –0.75 V (Fig. 2B, red). tion, to allow a more accurate comparison In contrast, the interior atom (Fig. 2B, blue) with the theoretical DOS (22). This proce- has an additional peak at þ0.5 V inside the dure maps spatial variations in the DOS band gap. along the chains (23). For example, the states Spectroscopy along the chains further re- seen in the individual spectra for the three- vealed quantized electronic states that are atom chain in Fig. 2D (blue) are mapped in confined within the isolated chain segments. Fig. 3D. The state at þ0.5 V (blue circles in An isolated atom displays a single state at Fig. 2D) appears as a dot that is localized to –0.75 V, which splits into two states at –0.75 the central atom (position 2 in Fig. 3D). In and –0.45 V for a two-atom dimer (Fig. 2, C contrast, the state at –0.7 V (blue triangles in and D). This split is the expected hybridiza- Fig. 2D) has the highest intensity over the tion for a dimer, although the splitting be- end atoms at positions 1 and 3, with lower tween the states is smaller than in previous intensity in the middle. States in the upper studies (9–11). To best distinguish the two corners (90.8 V) are localized to the regions

Fig. 2. On the left (A), an STM topography image shows a seven- atom chain with (B) selected spectra taken in the region be- yond the chain (green), over an end atom (red), and over an atom second from the end (blue). On the right (C), an STM topography image shows several chains ranging in length from one to four atoms. (D) Spectroscopy shows one state Fig. 1. The effects of end states are visible by for an isolated atom that splits comparing STM constant-current mode (0.1 nA) into two states for a dimer, as is images of the same area (8 by 19 nm) of Si(553)- expected from the atomic hy- Au at positive and negative sample biases. The bridization. The three-atom chain chains appear shorter at þ0.5-V sample bias (A) shows a new positive state that than at –1-V sample bias (B). The areas inside is much higher in energy and is the red boxes in (A) and (B) are expanded to localized over the middle atom. show an enlargement of a nine-atom chain in The image sizes are (A) 2 by 4 nm (C)and(D), respectively, and line profiles of the and (C) 2.5 by 5 nm. The set point topography along the chain are compared with voltages are –1.2 V for (A) and a tight-binding model in (E) and (F). The end (B) and –0.7 V for (C) and (D). atoms are barely visible at þ0.5 V [(C) and (E)] Spectroscopy curves are offset by 0.1 nS in (B) and 0.4 nS in (D) for ease of display and offset but are enhanced at –1 V [(D) and (F)]. baselines for each are also included.

704 4 FEBRUARY 2005 VOL 307 SCIENCE www.sciencemag.org R EPORTS beyond the chain ends and are attributable to persion relation (11). The n 0 1, 2, and 3 accumulation of intensity for the filled states either (i) the onset of the conduction band or quantum well states have evenly spaced en- in the range of –0.6 to –0.8 V. Furthermore, (ii) empty defect states. The apparent high ergies and exhibit one, two, and three peaks similar end states were formed for longer intensity beyond the ends of the chains is in the DOS along the chains, respectively. This chains (25). A tight-binding model with the misleading because of the marked inward result contrasts with the experimental data, same few parameters provides a consistent, motion of the tip (91 )) that leads to an which has only two resolved states with very semiquantitative description of the experi- exponential increase in the differential con- different topology (Fig. 3D). The empty states mental data for chain lengths ranging from ductivity (23). are missing intensity over the end atoms, four to nine atoms (26). If the model is ex- To interpret the DOS, we use a tight- whereas the filled states have high intensity tended to a semi-infinite chain, the parameters 0 binding model in which we consider the states over the end atoms. Assuming the total DOS we derived, with binding energy shift e1 – e0 in the chains as atomic wave functions, approx- at each atomic position is conserved, this re- –0.33 eV at the end atom that is comparable to 0 imated as a Gaussian at each site, interacting sult implies a transfer of DOS from the empty the hopping integral t1 –0.34 eV, are con- in one dimension. The hopping integral between to the filled states. Adding a large positive sistent with the formation of a localized end 0 the wave functions leads to the formation of second-nearest-neighbor hopping integral, t2 state (2). The inclusion of second-atom terms hybridized states that are a mixture of the 0.2 eV, to the model effectively lowers the in the binding energy shift suggests electronic atomic wave functions, similar to hybridized energy of the n 0 2 state so it is nearly de- end effects that extend at least two atoms into states in a molecule. The resulting chain wave generate with n 0 1 (Fig. 3B) but cannot ex- the chain. STM topography data from Fig. 1 functions and eigenenergies are parameterized plain the redistribution of DOS over the end reflect this assertion and show a clear en- in terms of the hopping integrals and the bind- atoms. For Fig. 3C, we used different bind- hancement of the second atom at positive ing energies at each atomic site, with the ing energies for the end and interior atoms bias that corresponds to the increased binding 0 hopping integrals determining the bandwidth of–0.6and0.27eV,respectively,witht1 energy of the second atoms in the model (26). 0 and the shape of the band for an infinite chain. –0.19 eV and t2 0.1 eV, which provides the Our spectroscopy measurements of finite The advantage of the tight-binding approach best agreement with the experimental data. chains also show how end effects can lower is that it allows differentiation of states at the The n 0 1 and n 0 2 states are nearly de- the energy of the filled states within the chains, end atoms. By shifting the electron binding generate and have enhanced DOS over the which suggests a possible driving force for energy on atoms at the ends of the chain rel- end atoms. The resulting redistribution of DOS the formation of end states. For the four-atom ative to the interior atoms, we can model the matches qualitatively the DOS in the exper- and five-atom chains, the lowest three states formation of end states. iment and explains why only two states are are nearly degenerate in energy, with the end A comparison of the DOS for a three- resolved (Fig. 3D). Thus, we can directly effects providing a substantial energy savings atom chain from the tight-binding calculation attribute the redistribution of DOS to the elec- (0.4 eV for the filled states in the five-atom (Fig.3,A,B,andC)andexperiment(Fig.3D) tronic end effects. In an alternative represen- chain) when compared with models in which provides the simplest demonstration of the tation, the transfer of DOS implies a transfer the end effects are omitted. Similar energy electronic end effects. First, consider the chain of charge from the interior to the end atoms. savings for the filled states are calculated for without including the end states and with only Spectroscopy of longer chains shows sim- all of the chains studied. In contrast, this mech- 0 the nearest-neighbor hopping integral, t1 ilar end states, as is seen for four- and five- anism is not applicable to previously studied –0.35 eV (Fig. 3A) (24). The electronic bind- atom chains (Fig. 3, E to H). In both cases, chain systems of Au on NiAl(110) and Cu on ing energy determines the energy zero, and the transfer of DOS over the end atoms is Cu(111), which have only empty states (9–11). the result is similar to a particle in a simple- similar to that for the three-atom chain, with The formation of end states does not lower the square well potential with a tight-binding dis- almost no intensity for the empty states and an energy unless they are filled, possibly explain- ing why a particle-in-a-box model was suf- ficient in these previous studies. The formation of end states provides a mechanism, in addi- tion to the Peierls distortion (27), to lower the total energy for 1D chains of finite length. From the calculated DOS in the chains, we can model the constant-current STM pro- files along the chains and thus verify the ori- gins of the end effects seen at different bias voltages in Fig. 1. Figure 1, E and F, com- pares calculated and experimental profiles along a nine-atom chain for tunneling into the empty and filled states. The model uses the calculated DOS with the aforementioned parameters (26) along with a transmission function that describes the decay of the wave functions into the vacuum. We further assume Fig. 3. Tight-binding calculations of the DOS and experimental differential conductance mea- surements as a function of bias voltage and position along the chain for three-, four- and five- a constant DOS for the tip and neglect tip atom chains. The magnitude of the differential conductance is represented on a rainbow color convolution effects. The calculated constant- scale, with red as high intensity and blue as low intensity, and plotted as a function of position on current profiles exhibit pronounced dips be- the x axis and sample bias on the y axis. For the three-atom chain, different tight-binding tween atoms that are much weaker in the parameters are tested (A to C) and are compared with an experimental dI/dV measurement taken experiment because of the finite size of the tip. with a constant-current set point (D). Only the model that includes end states (C) agrees with the Nevertheless, the relative heights at the atomic experiment. The presence of end states is confirmed by extending the model to (E) four-atom and (G) five-atom chains, providing good agreement with the experimental dI/dV measurements in (F) position (filled circles in Fig. 1, E and F) are in and (H), respectively. The distance along the chains is measured in units of the Si lattice spacing fair agreement with the experiment, especially along the chain direction (3.84 A˚). considering the simplicity of the model.

www.sciencemag.org SCIENCE VOL 307 4 FEBRUARY 2005 705 R EPORTS References and Notes 14. K. N. Altmann et al., Phys. Rev. B 64, 035406 (2001). 25. J. N. Crain, D. T. Pierce, data not shown. 1. A. Zangwill, Physics at Surfaces (University Press, 15. R. Losio et al., Phys. Rev. Lett. 86, 4632 (2001). 26. Consistency among different chain lengths requires Cambridge, 1988). 16. P. Segovia, D. Purdie, M. Hengsberger, Y. Baer, Nature the inclusion of (i) a nearest-neighbor hopping in- 0 2. S. G. Davison, M. Ste˛s´licka, Basic Theory of Surface 402, 504 (1999). tegral for the end atoms t1end –0.19 eV that is re- States (Oxford Univ. Press, New York, 1992). 17. An STS study of Si(111) 52-Au revealed 1D Schottky duced as compared with the nearest-neighbor hopping 0 3. M. V. Bollinger et al., Phys.Rev.Lett.87, 196803 barriers between insulating and metallic chain segments. integral between central atoms t1 –0.34 eV, (ii) a 0 (2001). 18. H. S. Yoon, S. J. Park, J. E. Lee, C. N. Whang, I. W. Lyo, second-nearest-neighbor hopping integral of t2 0 4. L. Burgi, O. Jeandupeux, A. Hirstein, H. Brune, K. Kern, Phys. Rev. Lett. 92, 096801 (2004). 0.10 eV, and (iii) binding energies of e1 –0.60 eV 0 Phys. Rev. Lett. 81, 5370 (1998). 19. J. N. Crain et al., Phys. Rev. Lett. 90, 176805 (2003). for the end atoms, e2 –0.09 eV for the atoms second 0 5. M. F. Crommie, C. P. Lutz, D. M. Eigler, Nature 363, 20. J. R. Ahn, H. W. Yeom, H. S. Yoon, I. W. Lyo, Phys. from the end, and e0 –0.27 eV for the remaining 524 (1993). Rev. Lett. 91, 196403 (2003). atoms in the interior of the chain. For the four-atom

6. Y. Hasegawa, P. Avouris, Phys. Rev. Lett. 71, 1071 21. H. W. Yeom et al., Phys. Rev. Lett. 82, 4898 (1999). chain there are no middle atoms, so only energies e1 (1993). 22. M. M. J. Bischoff, T. Yamada, A. J. Quinn, H. van Kempen, and e2 are used. 7. J. Repp, G. Meyer, K. H. Rieder, Phys. Rev. Lett. 92, Surf. Sci. 501, 155 (2002). 27. R. F. Peierls, Quantum Theory of Solids (Clarendon, 036803 (2004). 23. Two caveats must be considered in interpreting Oxford, 1955). 8. M. F. Crommie, C. P. Lutz, D. M. Eigler, Science 262, any spatially resolved STS measurements. The DOS 28. We thank M. D. Stiles for insightful discussions and 218 (1993). is convolved with the finite size of the tip leading suggestions concerning modeling, J. A. Stroscio for 9. N. Nilius, T. M. Wallis, W. Ho, Science 297, 1853 (2002). to a spatial broadening. Also, a constant-current helpful discussions and for building the STM, R. J. 10. T. M. Wallis, N. Nilius, W. Ho, Phys. Rev. Lett. 89, set point determines the tip-sample separation for Celotta and J. W. Gadzuk for helpful comments, 236802 (2002). each measurement, leading to an exponential in- and S. R. Blankenship for technical assistance. This 11. S. Folsch, P. Hyldgaard, R. Koch, K. H. Ploog, Phys. Rev. crease in the differential conductivity when the tip- work was supported in part by the Office of Naval Lett. 92, 056803 (2004). sample separation decreases, such as off the ends of Research. 12. J. N. Crain et al., Phys. Rev. B 69, 125401 (2004). the chains.

13.J.R.Ahn,H.W.Yeom,H.S.Yoon,I.W.Lyo,Phys. Rev. 24. Choices with positive t1 are ruled out from modeling 29 October 2004; accepted 23 December 2004 Lett. 91, 196403 (2003). longer chains. 10.1126/science.1106911

Here, we use carbon and sulfur isotopic Photic Zone Euxinia During data and biomarker and iron speciation analyses in a drill core (Hovea-3) from the the Permian-Triassic onshore Perth Basin, Western Australia (13), to establish the redox conditions in the water column of the southern Tethys Ocean Superanoxic Event during the Permian-Triassic (P-T) super- Kliti Grice,1* Changqun Cao,2 Gordon D. Love,3 anoxic event. Biomarkers diagnostic for an- Michael E. Bo¨ttcher,4 Richard J. Twitchett,5 oxygenic photosynthesis by Chlorobiaceae were identified in P-T boundary sediments Emmanuelle Grosjean,3 Roger E. Summons,3 6 1 2 of the organic-matter (OM)–rich Hovea-3 Steven C. Turgeon, William Dunning, Yugan Jin core and in coeval samples from the OM- lean Meishan-1, a new core drilled at the type Carbon and sulfur isotopic data, together with biomarker and iron speciation section of Meishan, South China (fig. S1); analyses of the Hovea-3 core that was drilled in the Perth Basin, Western these biomarkers demonstrate that waters Australia, indicate that euxinic conditions prevailed in the paleowater column of the Tethys Ocean were periodically eux- during the Permian-Triassic superanoxic event. Biomarkers diagnostic for inic in the photic zone during and after the anoxygenic photosynthesis by Chlorobiaceae are particularly abundant at the extinction event. boundary and into the Early Triassic. Similar conditions prevailed in the Changhsingian and Griesbachian sedi- contemporaneous seas off South China. Our evidence for widespread photic- ment samples of Hovea-3 (1960- to 1995-m zone euxinic conditions suggests that sulfide toxicity was a driver of the depth) contain C18 and C19 aryl isoprenoids extinction and a factor in the protracted recovery. (Figs. 1A and 2A), and the Griesbachian

sediments contain isorenieratane, the C40 The most severe extinction of the past 500 sphere provide a kill mechanism that might parent hydrocarbon (Figs. 1A and 2B and million years occurred in the Late Permian account for the terrestrial and marine extinc- fig. S2). Highly specific bacteriochlorophylls (1, 2). The biotic crisis was accompanied by tions (7). can also give rise to distinctive maleimides an oceanic anoxic event (OAE) that may In anoxic zones of modern-day stratified (9). Methyl iso-butyl maleimide was identi- have lasted up to 8 million years. Although lakes or restricted marine environments (e.g., fied in the polar fractions Esee (13)for different authors report various anoxic inter- the Black Sea and Antarctic fjords), con- separation of maleimides^. The highest con- vals, the most severe conditions persisted ditions are favorable for bacterial reduction centrations of all these pigment derivatives during the first 1 to 3 million years (3, 4). of sulfate to sulfide (e.g., 8). Chlorobiaceae are preserved in the Griesbachian, reflecting Anoxia has been proposed to have had a (green sulfur bacteria) are typical of these high green sulfur bacterial activity and, thus, major role in driving the extinction (5, 6); environments in which hydrogen sulfide PZE conditions. Isorenieratane and aryl surface outcropping of sulfidic waters and extends into the photic zone, where it serves isoprenoids (including low pristane/phytane emissions of hydrogen sulfide to the atmo- as the electron donor required for anoxy- ratios) also occur in the latest Changhsingian genic photosynthesis. Chlorobiaceae use a and earliest (Griesbachian) sediments 1Curtin University of Technology, Perth, Australia. distinct assemblage of light-harvesting pig- (beds 22 to 27) of the global boundary strato- 2Nanjing Institute of Geology and Palaeontology, ments comprising bacteriochlorophylls c, d, type section and point (GSSP) at Meishan, Nanjing, China. 3Massachusetts Institute of Technol- 4 and e and the carotenoids isorenieratene and South China, which suggests that PZE condi- ogy, Cambridge, MA 02139, USA. Max Planck chlorobactene. Identification of these com- tions were widespread (Fig. 1B and Fig. 3). Institute for Marine Microbiology, Bremen, Germany. 5Plymouth University, , UK. 6Oak Ridge pounds, or their diagenetic alteration pro- Although the above data provide evi- National Laboratory, Oak Ridge, TN 37831, USA. ducts, in sediments provides unequivocal dence for PZE during the P-T transition, the *To whom correspondence should be addressed. evidence for photic zone euxinic (PZE) con- presence of benthic epifaunal macroinverte- E-mail: [email protected] ditions in the past (e.g., 9–12). brates such as Claraia and spirorbids dem-

706 4 FEBRUARY 2005 VOL 307 SCIENCE www.sciencemag.org R EPORTS onstrates that, because these animals would phytane vary gradually across the P-T tran- C14-C18 n-alkyl carbon chains have many have required some oxygen, the euxinia was sition (Fig. 2F), representing in part a change inputs, comprising primary producers and episodic (14). in OM inputs or an increase in stratifica- heterotrophs, and their d13C values represent Independent evidence for euxinic con- tion toward the Triassic that caused enhanced a weighted average of these. If derived from 13 ditions in the Lower Triassic seas off Australia recycling of C-depleted CO2.However, primary sources such as algae and cyanobac- is provided by high amounts of metal- recent d13C data on higher plant and phyto- teria, they should be depleted in 13C com- loporphyrins, in particular vanadyl (VO) plankton biomarkers show similar isotopic pared with the co-occurring isoprenoids by porphyrins (Fig. 2C) (15). These samples also changes across the P-T transition, which 1.5° (e.g., 25, 26). This pattern is observed contained high concentrations of Ni (II) indicates a global disruption of the carbon in the Triassic data (Fig. 4), in which there is porphyrins. cycle (24). independent evidence, such as the abun- We also measured iron species, dithionite- The d13C values of pristane and phytane, dance of porphyrins and algal microfossils, extractable (FeD), pyrite (FeP), and total iron derived from phytol side chains of the for high primary productivity. Alternatively, þ (FeT)(16) in the Hovea-3 sediments. (FeD chlorophylls of algae and cyanobacteria, are isotopic enrichment of C14-C18 n-alkyl car- FeP)/FeT values up to 0.7 indicate euxinic robust proxies for the isotopic composition bon chains can occur through heterotrophic conditions in the basal-most Triassic (Fig. of phytoplankton (e.g., 25, 26). In contrast, processing of primary photosynthate or dom- 2D), comparable to values reported from the modern euxinic Black Sea (17). Variations in the ratio may be due to changes in the oxy- cline position, the area of sediment covered by anoxic waters, and changes in weathering rates. The isotopic composition of the oceanic sulfur reservoir is partly controlled by the balance of bacterial sulfate reduction and sulfide oxidation. These processes lead to 34S-depleted sulfide and related pyrite (18) and may result in sulfate relatively enriched in 34S. Burial of pyrite thus removes isotopi- cally light sulfur from the seawater pool. Sedimentary sulfides in the Hovea-3 samples (Fig. 2E) are depleted in 34S compared with contemporaneous seawater sulfate (19, 20)and follow a trend toward heavier values approach- ing the P-T transition, as seen in carbonate- associated sulfate in other P-T samples from northern Italy and Iran (19, 20). Sulfur-isotope fractionations between sedimentary pyrite and contemporary sulfate (19, 20) up to about 50 to 60 per mil (°) indicate that reservoir effects did not substan- tially influence the isotopic signatures. The 34S/32S ratios for the sulfides in the Basal Triassic of the Perth Basin are consistent with euxinic conditions as found in the modern Black Sea and the Pliocene Medi- terranean Basin (21). The change in isotope discrimination implies that there was a per- turbation in the sulfur cycle in the Upper Permianandarelativeincreaseinthe fraction of sulfur buried as pyrite in the Lower Triassic compared with the Permian (22) or that there was a change in other fac- tors (e.g., quality of OM) influencing overall sulfur isotope discrimination. Isotope shifts in reduced sulfur across the P-T boundary indicate changes in the sulfur cycle similar to those reported in sections from Japan (22). Stable carbon isotopic data from the bulk kerogen fraction of Hovea-3 record an abrupt Fig. 1. Typical gas chromatography–mass spectrometry selected-ion chromatogram for the 133-Da 7.5° negative shift from the Upper Per- fragment ion that is diagnostic for aryl isoprenoids of an aromatic hydrocarbon fraction [see (14)for mian to the Lower Triassic consistent with a fractionation of extracts]. (A) From 1979.9 m (Hovea-3). This trace shows pseudohomologous series C12- C compounds identified as aryl isoprenoids with a 2,3,6-trimethyl substitution pattern and the C localized palynofacies change from charcoal- 31 40 biomarker isorenieratane. These hydrocarbons were identified by comparison with a reference sample wood dominated OM to algal-amorphous (fig. S2). (B) From 107.94 m (Meishan-1). This trace for Meishan Bed 24-6 shows an assemblage of OM, respectively (23). In contrast, d13Cval- Chlorobiaceae biomarkers similar to those seen in Hovea-3. The enhanced relative abundance of ues of the molecular pristane and isorenieratane reflects a lower degree of thermal maturity in the sediments from South China.

www.sciencemag.org SCIENCE VOL 307 4 FEBRUARY 2005 707 R EPORTS

Fig. 2. Composite plot of Hovea-3 organic and inorganic data. (A)Aryl isoprenoid concen- trations (mg/g of to- tal organic carbon). (B) Isorenieratane con- centrations (mg/g of total organic carbon). (C) Concentrations of vanadyl and nickel por- phyrins (mg/g of total organic carbon) [see (14) for analysis of por-

phyrins]. (D)FeHR/FeT ratios. FeHR,highlyre- active iron; FeT,total iron; FeP, pyritic iron [see (14) for isolation of iron species]. (E) d34S of pyrite relative to Vienna Canyon Dia- blo troilite (VCDT) standard in ° [see (14) for sulfur isotope analyses]. (F) d13Cof pristane and phytane relative to Vienna Pee Dee belemnite (VPDB) standard in ° [see (14) for 5A molecular sieving and compound-specific isotope analyses].

Fig. 3. Composite plot of Meishan-1 organic geochemical, biomark- er, and bulk carbon isotopic data. Pristane/ phytane ratios show their lowest values just below and just above the extreme negative carbon-isotopic excur- sion. Maxima of aryl isoprenoid and iso- renieratane abundances in beds 24 to 27 bracket the extinction horizon. Notably, there are additional maxi- ma of these markers at beds 34, 35, and 37, which indicates that therewasawaxing and waning of PZE conditions that con- tinued well into the Triassic (fig. S1).

inant inputs of isotopically heavy bacterial and kerogen are similar to those seen at that similar conditions prevailed elsewhere biomass. This pattern of anomalous isotopic Meishan and the Southern Alps. Together, in the Tethys Ocean during the P-T event, we ordering is seen in the Permian sediments. these data provide further evidence for a propose that sulfide toxicity in the ocean and Figure 4 demonstrates this phenomenon, switch in the mode or extent of organic emissions of hydrogen sulfide to the atmo- which suggests that there was a shift in the carbon remineralization at the P-T transition. sphere were important drivers of the largest mode of carbon cycling at the P-T transition The data show that PZE conditions mass extinction in the past 500 million years in Hovea-3. A similar reversal in the occurred during the P-T superanoxic event and may have also been a factor in the expected C-isotopic relation between poly- and document a major disruption of the carbon protracted recovery (7). methylenic and polyisoprenoid lipids has and sulfur cycles. The onset of PZE in Hovea-3 The local association of PZE and high- been reported for the Proterozoic- coincides with a sharp facies change, re- sedimentary total organic carbon contents transition (27). Moreover, this transition in C- flecting rapid transgression (e.g., 23). The (TOC) at Hovea-3 are similar to those of other isotopic relations is also evident in data from association of oxygen-poor water and rapid Mesozoic OAEs (29). However, the con- the P-T transition of Western Slovenia (28), transgression is key for the hypothesis that ditions creating the record here were not where the isotopic trends of inorganic carbon anoxia caused the extinction event (5). Given repeated elsewhere, even within Western

708 4 FEBRUARY 2005 VOL 307 SCIENCE www.sciencemag.org R EPORTS Fig. 4. Dd 0 (average 18. D. E. Canfield, A. Teske, Nature 382, 127 (1996). 13 19. R. J. Newton, E. L. Pevitt, P. B. Wignall, S. H. Bottrell, d Cofn-C17 and n-C ) – (average d13C Earth Planet. Sci. Lett. 218, 331 (2004). 18 20. C. Korte et al., Int. J. Earth Sci. 93, 565 (2004). of pristane and phy- ° 21. H. F. Passier et al., Nature 397, 146 (1999). tane) (relative to 22. Y. Kajiwara, S. Yamakita, K. Ishida, H. Ishiga, A. Imia, VPDB standard in °) Palaeogeogr. Palaeoclimatol. Palaeoecol. 111, 367 with depth (meters). (1994). Measurements deter- 23. C. B. Foster, G. A. Logan, R. E. Summons, J. D. Gorter, mined by isotope- D. S. Edwards, Aust. Pet. Prod. Explor. Assoc. J. 37, ratio-monitoring gas 472 (1997). chromatography– 24. K. Grice et al., unpublished data. 25. J. M. Hayes, Rev. Mineral. Geochem. 43, 225 (2001). mass spectrometry 26. S. Schouten et al., Geochim. Cosmochim. Acta 62, (IsoPrime, Micromass, 1397 (1998). Manchester, UK) of 27. G. A. Logan, J. M. Hayes, G. B. Hieshima, R. E. Summons, saturate, branched/ Nature 376, 53 (1995). cyclic, and n-alkane 28. V. Schwab, J. E. Spangenburg, Appl. Geochem. 19,55 fractions [see (14) (2004). for 5A molecular siev- 29. R. D. Pancost et al., J. Geol. Soc. London 161, 353 ing and compound- (2004). 30. M. D. Lewan, J. B. Maynard, Geochim. Cosmochim. specific isotope analyses]. Acta 46, 2547 (1982). 31. Y. Shen, A. H. Knoll, M. R. Walter, Nature 423, 632 9. K. Grice et al., Geochim. Cosmochim. Acta 60, 3913 (2003). Australia, and must have been very localized. 32. We thank Origin Energy for collecting a unique, core- Because PZE also occurred in the contempo- (1996). 10. R. E. Summons, T. G. Powell, Geochim. Cosmochim. based maturity profile of the Hovea core of the Perth raneous seas off South China, localized high Acta 51, 557 (1987). Basin. We also thank S. Wang, G. Chidlow, and Geo- 11. M. Koopmans et al., Geochim. Cosmochim. Acta 60, science Australia (Canberra) for technical input and algal productivity probably played a key role M. Kuypers, D. Fike, and two anonymous reviewers for in the formation of a petroleum-rich source 4467 (1996). 12. K. Grice, P. Schaeffer, L. Schwark, J. R. Maxwell, comments. K.G. acknowledges the Australian Research rock in the Perth Basin. Org. Geochem. 25, 131 (1996). Council for funding. C.Q.C. and Y.G.J. were supported 13. Materials and methods are available as supporting by the China Ministry of Science and Technology and References and Notes material on Science Online. the National Natural Science Foundation of China. 1. D. Erwin, Nature 367, 231 (1994). 14. B. M. Thomas et al., Aust. J. Earth Sci. 51, 423 (2004). M.E.B. acknowledges Max-Planck-Gesellschaft for 2. M. J. Benton, R. J. Twitchett, Trends Ecol. Evol. 18, 15. Sedimentary porphyrins provide a link to precursor support. Work at the Massachusetts Institute of Tech- 358 (2003). chlorophylls in photosynthetic organisms indicative nology was supported by NASA Exobiology grant 3. P. B. Wignall, R. J. Twitchett, Science 272, 1155 (1996). of primary production in paleowaters. Ni/VO porphy- NAG5-1236. 4. Y. Isozaki, Science 276, 235 (1997). rin ratios are indicators of redox conditions (30). 0 þ 5. P. B. Wignall, A. Hallam, Palaeogeogr. Palaeoclima- 16. FeHR FeD FeP to FeT is an indicator of water- Supporting Online Material tol. Palaeoecol. 93, 21 (1992). column redox conditions in ancient/modern environ- www.sciencemag.org/cgi/content/full/1104323/DC1 9 6. A. H. Knoll, R. K. Bambach, D. E. Canfield, J. P. ments (17, 31). FeHR/FeT 0.4 are typical for euxinic Materials and Methods Grotzinger, Science 273, 452 (1996). settings with pyrite formation in the water column. Figs. S1 and S2 G 7. L. R. Kump, A. Pavlov, M. Arthur, Y. Kato, A. Riccardi, FeHR/FeT values 0.4 are found in sediments below References Abstracts of GSA Annual Meeting, Paper 81-6 oxic bottom waters, where pyrite precipitates in the G (Geological Society of America, Seattle, WA, 2–5 sediment. FeHR/FeT ratios 0.4 do not exclude 20 August 2004; accepted 6 January 2005 November 2003). deposition in a euxinic environment but may indicate Published online 20 January 2005; 8. T. et al., EOS Trans. Am. Geophys. Union 82, changes in the source/depositional setting (17). 10.1126/science.1104323 306 (2001). 17. T. F. Anderson, R. Raiswell, Am. J. Sci. 304, 203 (2004). Include this information when citing this paper.

synergistic combination of these (6), among Abrupt and Gradual Extinction Among others. An important test of any mechanism is a consideration of the pattern of extinc- Late Permian Land Vertebrates in the tions. The marine extinctions are well de- scribed in several areas, notably Meishan, China (7). Here, we report new chemostrati- Karoo Basin, South Africa graphic, biostratigraphic, and magnetostrati- Peter D. Ward,1* Jennifer Botha,3 Roger Buick,2 graphic data from multiple stratigraphic Michiel O. De Kock,5 Douglas H. Erwin,6 Geoffrey H. Garrison,2 sections located in the Karoo basin of South Africa that provide an exceptionally detailed Joseph L. Kirschvink,4 Roger Smith3 record of the terrestrial extinctions. Permian-Triassic (P-T) strata in the central The Karoo basin of South Africa exposes a succession of Upper Permian to Karoo basin provide the most intensively Lower Triassic terrestrial strata containing abundant terrestrial vertebrate investigated record of vertebrate fossils from fossils. Paleomagnetic/magnetostratigraphic and carbon-isotope data allow sections to be correlated across the basin. With this stratigraphy, the verte- 1Department of Biology, 2Department of Earth and brate fossil data show a gradual extinction in the Upper Permian punctuated Space Sciences, University of Washington, Seattle, by an enhanced extinction pulse at the Permian-Triassic boundary interval, WA 98195, USA. 3Karoo Paleontology, Iziko: South 4 particularly among the dicynodont therapsids, coinciding with negative African Museum, Cape Town, South Africa. Division of Geological and Planetary Sciences, California carbon-isotope anomalies. Institute of Technology, Pasadena, CA 91125, USA. 5Department of Geology, Rand Afrikaans University, Johannesburg, South Africa. 6Department of Paleo- The Permian extinction is universally por- ses include climate change due to increased biology, MRC-121 Smithsonian Institution, Washing- trayed as the most catastrophic of all atmospheric CO2 and/or CH4 (2), the effects ton, DC 20013, USA. Phanerozoic mass extinctions (1), yet its of extraterrestrial impact (3–5), the effects of *To whom correspondence should be addressed. cause remains problematic. Various hypothe- the eruption of the Siberian Traps, and some E-mail: [email protected]

www.sciencemag.org SCIENCE VOL 307 4 FEBRUARY 2005 709 R EPORTS the Upper Permian through the Triassic (8). identified a reversed polarity magnetozone Arctic. Although there is some uncertainty These strata are dominantly fluvial overbank (R1) È5 m beneath the Unit I-II contact about the reversal pattern near the P-T sediments deposited near the center of a (14). At Lootsberg and Wapadsberg, the boundary (16), recent records in Europe subsiding retroarc foreland basin (9). We have normal-polarity zone extends well up into suggest that the boundary occurs just above sampled across 200 m of the Palingkloof the (Unit IV). At Loots- the base of a reversed-to-normal-polarity Member of the and the berg, where the youngest strata were transition in Germany, although there has overlying Katberg Formation, where we sampled of all the sections, there is a change been a recent report placing it slightly lower, recognize four units spanning the Upper to reversed polarity (R2) above about 130 m in the uppermost part of the reversed chron Permian and Lower Triassic (10). Fossils and a final switch back to normal (N2) at the (17). The base of Unit II is thus approxi- were collected from these strata at five top. Given paleontological constraints (8), mately coincident with the P-T boundary. different areas. Hence, correlating the sec- we correlate the long normal found in Units These correlations were tested by mea- 13 tions, which is difficult between fluvial II, III, and most of IV with magnetozone suring sedimentary carbon isotope (d Ccarb 13 sections and has been notably problematic in SN1 of the classic German Trias sections and d Corg) stratigraphy at all the sections at the Karoo (11), is critical. (15) and thus define the top and bottom of meter or submeter sampling frequency (12) To correlate the stratigraphy, we obtained the Griesbachian stage in the Karoo. The (Fig. 2), a finer resolution than has been a magnetostratigraphic record (Fig. 1) (12) superjacent reversal at Lootsberg Pass is done before (18). This earlier P-T isotope from three sections Esome data are also in most parsimoniously correlated with magne- study in the Karoo found a single negative ^ 13 (13) . Samples from Unit II and Unit III are tozone SR1 of central Germany, R2 of the d Ccarb excursion approximately coincident all of normal polarity (Chron N1), and we Southern Alps, and GR1 of the Canadian with the last occurrence of the latest Permian

Fig. 1. Magnetostratigraphic correlation across the Karoo basin. The dashed lines represent the correlated positions between the stratigraphic sections.

710 4 FEBRUARY 2005 VOL 307 SCIENCE www.sciencemag.org R EPORTS zonal index fossil Dicynodon. The global d13C ues maintain a broad minimum in Unit II, a of these sections are extensively intruded by 13 record for the P-T boundary period is known pattern similar to global marine d Ccarb re- Mesozoic dolerite dikes and sills. We suggest in varying detail from several dozen marine cords. The lowermost negative excursion in that this igneous activity has homogenized the 13 13 and fewer nonmarine sites (e.g., 19, 20). In- d Ccarb observed in the Karoo most parsi- primary d Corg record at these two sections. tensively sampled sections show a gradual moniously correlates with the singular, lower Fossil vertebrate biostratigraphy confirms 13 12 13 decline in the sedimentary C: Cratio Griesbachian negative d Ccarb excursion that Unit II is essentially contemporaneous upward through the Upper Permian, then a reported recently from a Late Permian–Late across the Karoo basin. At all sections, the sudden decline of 92° at or near the pa- Triassic carbonate platform in the Nanpan- highest occurrence of the uppermost Permian leontologically defined extinction boundary, jiang Basin, Guizhou Province, China (22). zonal index, Dicynodon lacerticeps, is found 13 13 followed by a gradual increase in d C. The Furthermore, the broad swings in d Ccarb either immediately below (at the fossiliferous sudden d13C decline precedes the formal P-T values that were measured higher in the Karoo Lootsberg, Wapadsberg, and Bethulie sections) boundary based on the first occurrence of the sections (Fig. 2) appear to be consistent with or at most several meters below the base of 13 Triassic conodont Hindeodus parvus at the Smithian and Spathian age d Ccarb excur- Unit II (at the fossil-poor Carlton Heights 13 global stratotype section and point (GSSP) sions in the same marine d Ccarb record. and Kommandodrift Dam sections). Dicynodon P-T stratotype in Meishan, China, where the The bulk sedimentary organic carbon lacerticeps was never found in or above Unit 13 boundary has been placed at Bed 27. At isotope records (d Corg) from the Carlton II, and the first Triassic fossil common to all Meishan, the lowest d13C values are found in Heights and Lootsberg sections provide fur- sections— sp. C—was found in Bed 25 (7), coincident with the level of ther support for these conclusions (Fig. 2). The the lower strata of Unit III but never in Unit II. maximal species disappearance (94% loss of data also supports the magnetostratigraphic In summary, three independent correlation then-extant marine invertebrates). Thus, the correlation of Unit II between the northern methods support our contention that Unit II is mass extinction and sharp negative excursion (Carlton Heights and Bethulie regions) and both essentially isochronous across the Karoo in d13C are slightly older than the formal southern (Lootsberg and Wapadsberg re- basin and also time equivalent with the P-T stratigraphic boundary. gions) parts of the basin. At the Carlton boundary in China. This allows us to use this 13 We sampled both bulk sediment and Heights and Lootsberg sections, d Corg de- unit as a datum surface against which our carbonate paleosol nodules for carbon iso- creases from È –24° Vienna Pee Dee vertebrate range taxa can be plotted and to tope stratigraphy from our measured sec- belemnite (VPDB) to È –26° VPDB across compare the patterns of extinction with those 13 13 tions. Soil carbonate d C(d Ccarb)isa the uppermost meter of Unit I and remains observed at other P-T sections. function of both the d13C of atmospheric there through Unit II and into Unit III. Over a period of 7 years, we collected 13 13 CO2 (d CCO2) and the partial pressure of Between 15 m and 22 m (Unit III), d Corg 126 assigned to 21 vertebrate taxa È ° CO2 in the atmosphere (pCO2), and thus soil increases in both sections to –21 and from the sections shown here ( or 13 È ° d Ccarb records can be more scattered than then decreases 1 at the base of Unit IV. amphibian) (Fig. 3) (23). We treat these taxa 13 13 marine d Ccarb records (21). Nevertheless, This pattern is typical of P-T d C records as species, although further study will prob- 13 the d Ccarb results show a prominent drop measured elsewhere in the world. The ably result in an even greater number of taxa. 13 within Unit I that is consistent with our Wapadsberg and Bethulie d Corg records do We found 13 taxa in Unit I (Upper Permian), assessment that Unit I/Unit II contact marks not show any substantial negative excursions, only 4 of which persist into Unit II, and 12 13 the P-T extinction (Fig. 2). The d Ccarb val- particularly not at the P-T boundary, but both taxa in Units II to IV. Six of the 13 taxa

13 13 Fig. 2. The combined soil nodule d Ccarb record and individual lithologic Heights and Lootsberg Pass, however, have similar negative d Corg 13 13 and d Corg records for four sections in the Karoo basin. The two sections excursions within Unit II, followed by an increase in d Corgvalues in Units 13 at left (Wapadsburg, southern basin; Bethulie, northern basin) have been III and IV. The d Ccarb curve at the far right is combined data from strongly affected by heating, as evidenced by a lack of primary carbonate nodules obtained from the Carlton Heights and the Lootsberg paleomagnetic signal from either section. The results from Carlton regions.

www.sciencemag.org SCIENCE VOL 307 4 FEBRUARY 2005 711 R EPORTS found in Unit I show their last appearance Four taxa (Theriognathus sp., Dicynodon pect to observe, after the extinction, a follow- datums (LADs) in the last 10 m of Unit I, lacerticeps, Lystrosaurus sp. A, and Rubidgea on origination of new species into vacated which suggests an enhanced rate of extinc- sp.) appear to have become extinct at or near niches (or immigration into the basin). We do tion in the latest Permian. These range the base of Unit II (Fig. 3). We are also not observe this. By inverting the procedure terminations occurred either before or simul- confident that Pristerodon, Aelurognathus, for fossil disappearances, we calculated 20% 13 taneously with the reduced d Corg values and the abundant and widespread taxon range extensions downward and from this com- toward the top of Unit I. Diictodon sp. were extinct before deposition puted a 94% probability confidence interval for Incomplete preservation or collection fail- of Unit II, whereas the taxa Lystrosaurus sp. origination (Fig. 3). The results suggest it is ure can make abrupt extinctions look grad- B, Moschorhinus sp., and sp. probable that at least some species originated ual, i.e., the Signor-Lipps effect (24, 25), so became extinct during, or soon after, the ini- before the deposition of Unit II, whereas others the assumption of simultaneous extinction of tiation of sedimentation marking the Katberg originated during and after Unit II deposition. the included taxa was tested using the Formation (Unit IV) (Fig. 3). The 90% The distribution of originating taxa fails the Kolmogorov-Smirnov (K-S) goodness-of-fit confidence interval on the position of the K-S test for random distribution of new taxa at test (26). The case for the Karoo basin is mass extinction extends from 0 m (stippled the 99% level, suggesting that Triassic taxa complicated, because some of our sampled level A) to about 20 m, or from the base of origination was in response to some event that taxa (i.e., Pristerodon sp. and Aelurognathus Unit II up into the middle part of Unit III. occurred before the end of the Permian. We sp.) might have become extinct before the Thus, there is a low probability that the caution that new taxa origination may violate base of Unit II or long after (Ictidosuchoides mass extinction of the Karoo vertebrates the assumption of equal probability of preser- sp., sp., Lystrosaurus sp. C.), occurred within the Katberg Formation vation, because new taxa are likely to be rare thereby complicating the statistical protocols (stippled level B), where the mass extinction and uncommon. This effect may overestimate (27). Thus, we calculated confidence inter- has been traditionally placed (30). the calculated ranges for originations. vals on stratigraphic ranges (28). We chose We have also used confidence intervals A further caveat is that we are implicitly 20% confidence intervals following the im- to examine the origination of taxa. Theoret- accepting an empty-niche model brought proved method for testing extinction levels ically, if the majority of Permian extinctions about by the extinction, even though some (29) for each of the nine taxa (Table 1). occurred at a specific horizon, we would ex- or many niches may have been reconstructed

Fig. 3. Magnetostratigraphic, biostratigraphic, and carbon isotopic records the highest occurrence of Permian index fossil Dicynodon lacerticeps.Level from the Karoo basin. This figure can be compared with the generic range B is the base of Unit IV (Katberg Formation), where a spike in fungal spore chart for the entire Permian/Lower Triassic sequence in the study area (12). abundance and the P-T boundary have been recorded previously (32). Of The red bar on the left is the 90% confidence interval for Permian taxa particular note is the appearance of Lystrosaur AandLystrosaur B below the extinction; the red bar on the right is the 94% confidence interval for P-T boundary. Lystrosaurs are Triassic animals, and the appearance of two Triassic taxa origination, assuming that originations were in response to species below the P-T boundary is evidence that major evolutionary catastrophic extinction. Level A is the base of Unit II, which is correlated to changes were under way before the end of the Permian.

712 4 FEBRUARY 2005 VOL 307 SCIENCE www.sciencemag.org R EPORTS and not refilled. The pattern of extinction origination or appearance (through emigra- interval of even greater perturbation, i.e., a and origination appears to be consistent with tion) of new Triassic species into the basin. long Bpress[ with one (or more) pulses su- an enhanced rate of extinction coincident Our statistical inference (and the discovery of perimposed. A single proximal cause might with or just below Unit II, but the number of Lystrosaurus sp. A and B) that Triassic ver- explain the extinction patterns, such as long- extinct species there may be no more than tebrate fauna may have predated the main term environmental degradation having five (eight species become extinct in Unit I). Permian extinction pulse is unlike the pattern reached a critical threshold that triggered a The most thorough compilation of the ver- of mammalian radiation after the - short-term extinction event through eco- tebrate record from the Dicynodon Zone in Paleogene (K-P) extinction, the only clear system collapse. Alternatively, the long- the Karoo (8) shows that 10 of 21 taxa example of a mass extinction associated with and short-term effects observed in the Karoo present at the base of the È300-m-thick zone a major impact event. The pattern that we could have two (or more) different causes. are absent in its upper third, indicating that observe for the P-T is consistent with a long- We searched for evidence of an end-Permian considerable extinction is occurring through- term deterioration of the terrestrial ecosystem, bolide impact, such as the impact clays and out the zone and not just at its top (31). This with a heightened pulse of both extinction ejecta layers found commonly in the envi- observation can be supported by replotting and origination approximately coincident with ronmentally similar Hell Creek Formation known vertebrate ranges in the Karoo such the P-T boundary. (Late Cretaceous), Montana, which are that last occurrences are sequential (12), Unfortunately, the ranges of Permian and associated with the Chicxulub K-P impact. based on the new data presented here and Triassic fossil plant remains in our study Neither our search nor previous searches in range data from lower zones (8). The result- sections add little information about the the Karoo sections for minerals associated ing figure (fig. S4) shows what appears to be pattern of extinction in the Karoo basin or with large-body impact (34) have met with a change from an approximately constant about the relative timing of extinction among success, although fluvial facies can contain background extinction (as recognized by the plants and animals there. A recent palyno- hiatuses and the absence of impact evidence number of taxa disappearing per unit thick- logical study at Carlton Heights (32) identi- must be tempered with caution. ness of strata) below the uppermost Permian, fied a spike in fungal spore abundance at the The P-T southern Karoo basin and the K-P with enhanced extinction in the Dicynodon base of our Unit IV (the Katberg Formation) Hell Creek Formation strata were deposited Zone culminating at the P-T extinction pulse and claimed the fungal spike to be the top of by similar fluvial systems, and they can be at the top of Unit I. In the Lower Triassic– the Dicynodon Zone and thus the P-T bound- compared. The Hell Creek vertebrate paleon- aged Lystrosaurus Zone, there is a reduced ary. However, the study provided insufficient tological record is constantly diverse up extinction rate high in Unit II and in Unit III. information to ascertain whether the base of through the last Cretaceous zone, followed Two patterns are thus apparent: gradual Unit IV is Permian or Triassic in age based by a catastrophic extinction coeval with a extinction leading up to the P-T and a pulse on palynomorphs (33), and we have found negative excursion in the d13C record (35, 36) of even higher extinction marking the in all of our sections, including the site in and clear sedimentological and mineralogical boundary. This pattern is visible both at the question, that the Katburg Formation begins evidence of a large-body impact; d13C returns species level (Fig. 3) and at the generic level well above the top of the Permian. to pre-event values within a narrow strati- (fig. S4) and is also observed at Meishan We interpret the biostratigraphic data graphic interval (35). The Karoo record is among invertebrate marine fossils. presented here as consistent with a period entirely different, and we conclude that if an We thus suggest that factors other than of environmental stress during the latest impact occurred at all, it had a minor role in the sudden extinction of taxa stimulated the Permian in this basin, punctuated by a short the end-Permian extinctions in the Karoo. The geologic data from the Karoo are consistent Table 1. Confidence that vertebrates became extinct or originated prior to deposition of lithological Unit with a more protracted catastrophic ecosystem II by application of the equation C 0 1 – (G/R þ 1) –(H – 1), where G is the interval between the highest collapse than a sudden impact would produce. occurrence of the taxon and the base of Unit II, R is the taxon’s observed stratigraphic range, and H is the number of fossiliferous strata within the range of R (28). The null hypothesis of a random distribution of References and Notes fossil horizons is rejected. Taxon confidence (C), extinction before deposition of Unit II. Confidence, 1. D. H. Erwin, Nature 367, 231 (1994). origination before deposition of Unit II. 2. A. Knoll et al., Science 273, 452 (1996). 3. L. Becker et al., Science 291, 1530 (2001). 4. A. R. Basu et al., Science 302, 1388 (2003). Confidence (C) (extinction Confidence (origination Taxon 5. L. Becker et al., Science 304, 1469 (2004). before deposition of Unit II) before deposition of Unit II) 6. M. J. Benton, R. J. Twitchett, Trends Ecol. Evol. 18, 358 (2003). Pristerodon sp. 0.875 7. Y. G. Jin et al., Science 289, 432 (2000). Aelurognathus sp. 0.5 8. B. Rubidge, Geol. Surv. S. Afr. Biostratigraphy 1, (1995). Diictodon sp. 0.52 9. O. Catuneanu et al., Basin Res. 10, 417 (1998). Theriognathus sp. 0.31 10. Two sections were sampled near Lootsberg Pass Rubidgea sp. 0.2 (S31, 51.005; W24, 52.299, and S31, 49.334; W24, Dicynodon lacerticeps 0.17 48.565), one section near Wapadsberg Pass (S31, 52.474; W24, 54.882), one section near Carlton Lystrosaurus sp. A 0.13 Heights (S30, 35.425; W25, 439.135), one section Lystrosaurus sp. B 0 near Kommandodrift Dam (S31, 76.506; W24, 49.980), Pelanomodon sp. .05 and two sections near Bethulie (S30, 24.989; W26, Moschorhinus sp. 0 17.234, and S30, 26.675; W26, 18.006). Four litho- Owenetta sp. 0 stratigraphic facies are present: Unit I, dark gray to Ictidosuchoides sp. 0 gray mudstones, siltstones, and sandstones with sp. 0.08 sedimentary structures typical of meandering river sp. 0.2 deposits; strata show rubification in the uppermost meters. Unit II, 3- to 5-m-thick, rhythmically bedded sp. 0.25 laminated mudrock, described as an event bed (37). sp. 0.21 Unit III, red concretionary mudstone and thin Lystrosaurus sp. C 0.69 sandstone. Unit IV (Katberg Formation), thick olive- Micropholis sp. 0.08 green sandstone with conglomeratic bases interbedded Thrinaxodon sp. 0.55 with thinner red siltsone and mudstone; sandstones Procolophon sp. 0.54 have sedimentary structures typical of braided river deposits.

www.sciencemag.org SCIENCE VOL 307 4 FEBRUARY 2005 713 R EPORTS

11. G. Retallack et al., Geol. Soc. Am. Bull. 115,1133 taxonomic study of each is required. Pending the 34. L. Coney et al., L.P.S.C. 35, 1488 (2004). (2003). formal systematic treatment of Lystrosaurus (38), we 35. N. C. Arens, A. H. Jahren, Palaios 15, 4 (2000). 12. Materials and methods are available as supporting designate the four separate species of Lystrosaurus 36. P. Sheehan et al., Science 254, 835 (1991). material on Science Online. from our study area as Lystrosaurus sp. A, B, C, and D. 37. P. D. Ward et al., Science 289, 1740 (2000). 13. M. O. de Kock, J. L. Kirschvink, Gond. Res. 7, 175 (2004). 24. P. W. Signor III, J. H. Lipps, Geol. Soc. Am. Spec. Pap. 38. J. Botha, R. Smith, in preparation. 14. All samples passed tests for baked contact, class B 190, 291 (1982). 39. R. J. Twitchett et al., Geology 29, 351 (2001). reversal, and magnetostratigraphic consistency. The 25. Preservational biases may also have controlled the 40. C. Foster, M. Stephenson, First Int. Conf. Palynology, reversal in the upper part of Lootsberg Pass was observed pattern of extinction. However, because Abst. 57 (2002). corroborated at a second parallel section È1kmto there are more fossils in Units III and IV than in the 41. We thank the NASA Astrobiology Institute, the NSF, the east, where a reversal of similar thickness was upper 30 m of Unit I, we would expect to find the and the National Research Foundation of South found at the same stratigraphic horizon. We have Permian taxa if they continued higher in the section. Africa for funding. Help in the field and fossil prep- used this pattern to correlate and subdivide the As that is not the case, we conclude that the ob- aration came from the Karoo Paleontology Depart- Katberg Formation across the Karoo basin. served ranges are real samples of the preservable ment, Iziko: South African Museum (P. October, 15. M. Szurlies et al., Earth Planet. Sci. Lett. 212, 263 (2003). fauna in the depositional basin. H. Stumer, G. Farrell, preparation by A. Crean, field 16. M. Steiner et al., J. Geophys. Res. 94, 7343 (1989). 26. M. S. Springer, Paleobiology 16, 512 (1990). collection by N. Ward and T. Evans, and lab help by 17. J. Nawrocki, Terra Nova 16, 139 (2004). 27. A. Solow, Paleobiology 29, 181 (2003). C. Converse and E. Steig). Paleomagnetic software 18. K. G. MacLeod et al., Geology 24, 227 (2000). 28. C. R. Marshall, Paleobiology 20, 459 (1994). used for data analysis was from C. Jones at the 19. M. J. De Wit et al., J. Geol. 110, 227 (2002). 29. S. Wang, C. R. Marshall, Paleobiology 30, 5 (2004). University of Colorado, Boulder. We thank F. Kyte 20. W. T. Holser, M. Margaritz, Geochim. Cosmochim. 30. G. H. Groenwald, J. W. Kitching, Geol. Surv. S. Afr. and C. Looy for prereviews. Acta 56, 3297 (1992). Biostratigraphy 1, 35 (1995). 21. T. E. Cerling, Global Biogeochem. Cycles 6, 307 (1992). 31. G. M. King, S. Afr. Mus. Sidney Haughton Mem. Lect. Supporting Online Material 22. J. L. Payne, Science 305, 506 (2004). 3, 1 (1990). www.sciencemag.org/cgi/content/full/1107068/DC1 23. The actual P-T boundary is defined by the base of the 32. M. B. Steiner et al., Palaeogeogr. Palaeoclimatol. Materials and Methods Triassic system or the first appearance of the Palaeoecol. 194, 405 (2003). Figs. S1 to S4 conodont H. parvus in marine strata. The base of 33. There has been much recent criticism of the so- Tables S1 and S2 the Triassic cannot be identified in the Karoo until a called P-T fungal spike as a chronostratigraphic References terrestrial index fossil is formally chosen. At present, marker, based on the discovery of multiple horizons we have placed the P-T boundary at the level of the at some localities and the complete absence of the 3 November 2004; accepted 3 January 2005 highest Permian taxon, a practice that runs contrary marker at others, most notably at the important Published online 20 January 2005; to accepted stratigraphic procedure. Here, each Greenland locality (39), as well as the possibility that 10.1126/science.1107068 taxon is treated as a species; we realize that the fossils may not have come from fungi at all (40). Include this information when citing this paper.

kgd1D and kgd2D cells, less so in the pda1D, Aconitase Couples Metabolic pdb1D, idh1D,andlpd1D strains, and very unstable in aco1D cells(Fig.1A).mtDNA Regulation to Mitochondrial was previously noted to be unstable when some of these genes were inactivated (11). Further experiments established that ACO1 DNA Maintenance is essential for mtDNA maintenance. South- Xin Jie Chen, Xiaowen Wang, Brett A. Kaufman,* ern blot analysis of total cellular DNA from Ronald A. Butow. nascent meiotic segregants derived from an aco1D/ACO1 rþ diploid strain showed that Mitochondrial DNA (mtDNA) is essential for cells to maintain respiratory the aco1D spores lack mtDNA (Fig. 1B). competency and is inherited as a protein-DNA complex called the nucleoid. Aco1p, citrate synthase (CS), and two subunits of nicotinamide adenine dinucleotide We have identified 22 mtDNA-associated proteins in yeast, among which is þ mitochondrial aconitase (Aco1p). We show that this Krebs-cycle enzyme is (NAD )–dependent isocitrate dehydrogenase essential for mtDNA maintenance independent of its catalytic activity. Reg- (collectively, the aconitase metabolon) func- ulation of ACO1 expression by the HAP and retrograde metabolic signaling tion to produce a-ketoglutarate, a precursor pathways directly affects mtDNA maintenance. When constitutively ex- to glutamate. Expression of the genes encod- pressed, Aco1p can replace the mtDNA packaging function of the high-mobility- ing these proteins is positively regulated by group protein Abf2p. Thus, Aco1p may integrate metabolic signals and mtDNA the glucose-repressible HAP2-5 transcription maintenance. complex and the transcription factors Rtg1p and Rtg3p (12), which are components of the Mitochondrial DNA (mtDNA) nucleoids can be grouped into four functional catego- mitochondria-to-nucleus retrograde (RTG) sig- have been purified from several organisms ries: (I) mtDNA transactions with no other naling pathway (13). Aco1p localizes to the (1–5). In addition to DNA packaging pro- known functions in mitochondria; (II) protein mitochondrial matrix, as revealed by fluores- teins, which are required for mtDNA mainte- import and mitochondrial biogenesis; (III) cence microscopy of an Aco1p–green fluores- nance (6, 7), nucleoids also contain proteins the citric acid cycle and upstream glycolytic cent protein fusion construct (fig. S1). Like whose functions are ostensibly unrelated to steps; and (IV) amino acid metabolism. other genes in the aconitase metabolon, expres- mtDNA activities (1, 5, 8, 9). We previously As proteins in category III could poten- sion of ACO1 becomes progressively HAP- identified 11 proteins that are associated tially connect respiratory and fermentative dependent in response to increased respiratory with mtDNA (1) (Table 1) and have now metabolism to mtDNA maintenance, we ex- activity when cells are shifted from glucose identified 11 more (whose gene names are amined mtDNA stability in strains in which to raffinose medium, whereas a combined in- indicated in bold in Table 1). These proteins a selection of these genes were inactivated. activation of the HAP and RTG systems leads Expression of some category III genes is to a 14-fold reduction of ACO1 expression, Department of Molecular Biology, University of Texas repressed by glucose (10). Therefore, mutant independent of the carbon source (fig. S2). Southwestern Medical Center, 6000 Harry Hines cells were grown in raffinose, a fermentable To ask whether the HAP and RTG Boulevard, Dallas, TX 75390–9148, USA. carbon source that does not repress mitochon- transcription complexes can directly affect *Present address: Neurological Institute, drial respiration, and then assayed for the frac- mtDNA maintenance, we generated a diploid 3801 University, Montreal QC H3A 2B4, Canada. .To whom correspondence should be addressed. tion of respiratory-deficient (petite) mutants in strain heterozygous for hap2D and rtg1D. E-mail: [email protected] the population. mtDNA is relatively stable in After sporulation and dissection on rich

714 4 FEBRUARY 2005 VOL 307 SCIENCE www.sciencemag.org R EPORTS Fig. 1. ACO1 is required for mtDNA mainte- nance. (A) mtDNA stability in cells with null mutations of genes that encode metabolic pro- teins found in nucleoids. mtDNA stability is ex- pressed as percentage of respiratory-competent (rþ) colonies after growth for 35 generations in complete raffinose medium. Data are shown as means T SEM for triplicate experiments. WT, wild type. (B) Southern blot analysis of Cfo I–digested total DNA probed for mtDNA in nascent meiotic segregants containing ACO1 (þ)oraco1D (–) alleles. Controls are rþ and r0 cells. i.c., internal control for sample loading. (C) Southern blot analysis showing the loss of mtDNA in hap2D rtg1D meiotic segregants. (D) Suppression of mtDNA loss from the hap2D rtg1D double mutants by constitutive expression of a chromosomally integrated copy of ADH1- ACO1 (ACO1*).

glucose medium, both the rtg1D and hap2D spores can maintain mtDNA (Fig. 1C), whereas mtDNA was barely detectable in rtg1D hap2D segregants. However, when the diploids contained an integrated single copy of ACO1 under the control of the constitutive ADH1 promoter, the loss of mtDNA from the rtg1D hap2D double mutants was largely reversed in meiotic segregants that also received the ADH1-ACO1 allele (Fig. 1D). To determine whether the mtDNA in- stability in aco1D cells is due to a block in metabolic flux through the aconitase me- tabolon, we inactivated all three known genes encoding CS: CIT1, CIT2,andCIT3 encod- ing, respectively, the citric acid cycle CS, a peroxisomal isoform of CS, and a mitochon- drial CS that may function in a methyl citrate pathway (14, 15), so that no substrate would be available to aconitase. This resulted in a strong glutamate auxotrophy (Fig. 2A), dem- onstrating that metabolic flux through the aconitase metabolon had been blocked. Fig. 2. Functional bisection of Aco1p. (A) Growth of wild-type and mutant cells with the indicated Nevertheless, mtDNA was stable in the cit1D, citD alleles on minimal glucose medium with or without 0.2% glutamate (YNBD, 0.67% yeast cit2D, and cit3D triple mutant (Fig. 2B). nitrogen base, 2% dextrose). (B) Southern blot analysis of mtDNA in meiotic segregants from CIT/citD We next asked whether aconitase catalyt- diploid cells. (C) Schematic for generation of diploid strains used for the functional test of the ic activity is required for Aco1p function in cysteine mutants of ACO1. locations are indicated (XII and XV). (D) Southern analysis C448S C382S C445S mtDNA maintenance. Aconitase contains an showing that the aco1 but not the aco1 and aco1 alleles retain mtDNA among the meiotic segregants in a representative tetrad. iron-sulfur center (4Fe-4S) in which 3 of the 4 mol of iron are coordinated with three cys- teines, Cys382, Cys445, and Cys448 (16); the center is essential for aconitase enzymatic the mutants to see whether they could still fourth iron is coordinated with the substrate, activity (18). We changed each of these function in mtDNA maintenance. Each citrate (17). The integrity of this iron sulfur cysteine residues to a serine and analyzed mutant allele tagged with URA3 was inserted

www.sciencemag.org SCIENCE VOL 307 4 FEBRUARY 2005 715 R EPORTS into the ADE2 locus of an ACO1/aco1D::KAN (20). Accordingly, when HAP4 was expressed To show directly that the suppression of diploid strain (Fig. 2C). After sporulation, from the constitutive PGK1 promoter, the loss mtDNA instability of abf2D cells grown on Southern blot analysis of meiotic segregants of mtDNA from abf2D cells grown on glu- glucose is related to ACO1 expression, we ex- from a representative tetrad from each diploid cose was considerably delayed (Fig. 3C). pressed either the wild-type or one of the three strain showed that mtDNA was retained in Next, we inactivated the MKS1 gene cysteine mutants of Aco1p under control of the aco1D aco1C448S but not in the aco1D aco1C382S encoding a negative regulator of the RTG constitutive ADH1 promoter in centromeric plas- or aco1D aco1C445S segregants (Fig. 2D); how- pathway (21–23); this results in constitutive, mids. The transformants, maintained on glycer- ever, all three of the segregants are glutamate high levels of expression of RTG target ol medium, were examined for mtDNA stability auxotrophs and respiratory deficient, indicat- genes, including Aco1p. The mks1D muta- after plating on glucose medium. As expected, ing that they lack aconitase activity. These tion almost completely suppressed the loss of most abf2D cells transformed with vector alone differences in the ability of the aco1cys mu- mtDNA from abf2D cells grown on glucose formed petite colonies, indicating instability tant alleles to maintain mtDNA are not due (Fig. 3D). That the mks1D suppressor activ- of mtDNA (Fig. 3E). However, in cells express- to any appreciable differences in the expres- ity acts through the RTG pathway is shown ing wild-type Aco1p or its cysteine mutants, sion of the mutant proteins, because all are by the reappearance of mtDNA instability in the mtDNA instability phenotype was reversed, expressed at levels comparable to wild-type the abf2D mks1D double mutant when RTG1 yielding petite frequencies comparable to that Aco1p (fig. S3). Further experiments showed, or RTG3 was also inactivated. of the control cells expressing ABF2. however, that the aco1C382S and aco1C445S mutant alleles can support mtDNA mainte- nance when overexpressed from the strong ADH1 promoter, although those mutants re- main respiratory-deficient and auxotrophic for glutamate (fig. S4). The requirement of Aco1p for mtDNA maintenance is comparable to that of the mtDNA packaging protein Abf2p, which is essential for mtDNA maintenance in cells grown on glucose medium (6). The only conditions known in which mtDNA in abf2D cells is relatively stable is when another DNA packaging protein, such as Escherichia coli HU, is expressed in abf2D cells and targeted to mitochondria (19)orwhenabf2D cells are grown on a nonfermentable carbon source (6). This latter result has generally been in- terpreted to mean that abf2D cells can be propagated on medium such as glycerol be- cause of the strong selection for cells that retain mtDNA. However, under these growth conditions, ACO1 expression is elevated (10). This raises the possibility that, under dere- pressed conditions, Aco1p might be able to substitute for Abf2p in mtDNA maintenance. To test this, we first examined mtDNA sta- bility in abf2D cells grown on raffinose, which does not repress the aconitase me- tabolon. These experiments (Fig. 3A) show that growth on raffinose almost completely suppresses the mtDNA instability seen when abf2D cells are grown on glucose. Southern blot analysis of abf2D cells grown on glu- cose versus raffinose medium showed that after growth of the abf2D mutant in glucose for six generations, mtDNA was barely de- tectable in the cell population, whereas cells Fig. 3. Control of mtDNA inheritance by the HAP and RTG pathways. (A) mtDNA stability of abf2D grown in raffinose for the same number of þ generations retained a much greater amount cells grown in raffinose (Raf) or glucose (Glu) medium expressed as the percentage of r colonies on complete glucose medium after a growth in liquid medium for the indicated generations. (B) Southern of mtDNA (Fig. 3B). blot analysis of mtDNA in abf2D cells grown for six generations in complete glucose or raffinose We next determined whether the HAP- medium. Lane 1, rþ control; lane 2, abf2D;lane3,ro control; i.c., internal control. (C) Suppression of RTG targets are responsible for the marked mtDNA instability in abf2D cells by overexpression of HAP4 from the PGK1 promoter in rich glucose mtDNA stability observed in abf2D cells medium. abf2D cells were also transformed with an empty vector and with wild-type ABF2 on a grown under derepressed conditions. We first plasmid as controls. (D) Suppression of mtDNA instability in abf2D cells by constitutive activation of manipulated the HAP2-5 transcription com- the RTG pathway because of the mks1D mutation. (E) Suppression of mtDNA loss from abf2D cells by overexpression of ACO1 and its C382S, C445S, and C448S variants from the constitutive ADH1 plex by constitutively expressing HAP4,which promoter. mtDNA stability was determined by counting the fraction of rþ colonies after plating activates the expression of respiratory genes, transformants on complete glucose medium. (F) EB sensitivity of abf2D cells with ABF2, ACO1,oran including ACO1, in glucose-repressed cells empty vector on complete ethanol medium.

716 4 FEBRUARY 2005 VOL 307 SCIENCE www.sciencemag.org R EPORTS One interpretation for the suppression of of abf2D cells was also observed under glucose glucose repressed conditions, Abf2p is es- mtDNA instability in abf2D cells by activation repressed conditions (fig. S6). Together, these sential for mtDNA maintenance because of of ACO1 is that mtDNA is repackaged into data suggest that Aco1p may be providing its DNA packaging function. In derepressed more stable nucleoid structures. To investigate some packaging function for mtDNA. cells with robust oxidative metabolism, or in this, we measured the sensitivity of rþ cells to Our data establish a direct link between response to RTG signals, mtDNA packaging ethidium bromide (EB), which, by intercalat- control of ACO1 expression by the HAP and may also involve Aco1p. This raises the ing between the base pairs of mtDNA, RTG pathways and the maintenance of possibility that nucleoid remodeling may be produces petites because of the loss of the mtDNA. Our finding that Aco1p is required part of a strategy for adjusting mtDNA rþ mtDNA. EB accessibility to DNA has been for mtDNA stability independent of aconi- maintenance to the changes in cellular shown to be a useful probe for detecting tase catalytic activity is reminiscent of the metabolism. Because aconitase is susceptible differences in the organization of DNA-protein bifunctionality of the cytosolic form of to oxidative damage during oxidative stress complexes (24). Indeed, by comparison to mammalian aconitase, also known as iron- and cell aging (27–29), this property could wild-type rþ cells, abf2D cells are hyper- responsive element binding protein. That contribute to compromised mtDNA integrity sensitive to EB as determined by growth on protein switches between an enzymatic and as a result of a decoupling between metab- ethanol medium (Fig. 3F). This EB hyper- RNA binding form on the basis of the olism and mtDNA transactions. sensitivity can be suppressed by overexpres- assembly or disassembly of the E4Fe-4S^ sion of ACO1. We confirmed that, under these cluster (25, 26). Whether a similar mecha- References and Notes growth conditions, expression of ACO1 from nism applies to Aco1p function in mtDNA 1. B. A. Kaufman et al., Proc. Natl. Acad. Sci. U.S.A. 97, the multicopy plasmid increases the steady- maintenance remains to be determined. Our 7772 (2000). state level of Aco1p by threefold compared results further suggest that mtDNA nucleoids 2. S. Meeusen et al., J. Cell Biol. 145, 291 (1999). 3. I. Miyakawa, N. Sando, K. Kawano, S. Nakamura, T. with that in cells containing the empty vector may exist in different states depending on Kuroiwa, J. Cell Sci. 88, 431 (1987). (fig. S5). Aco1p suppression of EB sensitivity the metabolic condition of cells. Under 4. N. Garrido et al., Mol. Biol. Cell 14, 1583 (2003). 5. D. F. Bogenhagen, Y. Wang, E. L. Shen, R. Kobayashi, Mol. Cell. Proteomics 2, 1205 (2003). 6. J. F. Diffley, B. Stillman, Proc. Natl. Acad. Sci. U.S.A. Table 1. Mitochondrial nucleoid proteins identified by in organello formaldehyde cross-linking 88, 7864 (1991). (supporting online text). ATP, adenosine triphosphate KGDC, a-ketoglutarate dehydrogenase complex; 7. N. G. Larsson et al., Nature Genet. 18, 231 (1998). PHDC, pyruvate dehydrogenase complex; BCADC, branched-chain amino acids dehydrogenase complex. 8. B. A. Kaufman, J. E. Kolesar, P. S. Perlman, R. A. Genes encoding proteins identified in this study are boldfaced. Butow, J. Cell Biol. 163, 457 (2003). 9.H.Sato,A.Tachifuji,M.Tamura,I.Miyakawa,Proto- plasma 219, 51 (2002). Gene Protein Primary functions 10. J. L. DeRisi, V. R. Iyer, P. O. Brown, Science 278,680 (1997). Category I 11. M. T. McCammon, C. B. Epstein, B. Przybyla-Zawislak, ABF2 Abf2p mtDNA packaging, L. McAlister-Henn, R. A. Butow, Mol. Biol. Cell 14, rþ genome maintenance 958 (2003). MGM101 Mgm101p Maintenance of rþ and ori-less 12. Z. Liu, R. A. Butow, Mol. Cell. Biol. 19, 6720 (1999). mtDNA, mtDNA repair 13. R. A. Butow, N. G. Avadhani, Mol. Cell 14, 1 (2004). RIM1 Single-stranded DNA binding protein mtDNA replication 14. Y. K. Jia, A. M. Becam, C. J. Herbert, Mol. Microbiol. 24, 53 (1997). RPO41 DNA-directed RNA polymerase Mitochondrial transcription, þ 15. C. B. Epstein et al., Mol. Biol. Cell 12, 297 (2001). r mtDNA maintenance 16. A. H. Robbins, C. D. Stout, Proc. Natl. Acad. Sci. SLS1 Sls1p Mitochondrial translation, U.S.A. 86, 3639 (1989). þ r mtDNA maintenance 17. H. Lauble, M. C. Kennedy, H. Beinert, C. D. Stout, Category II J. Mol. Biol. 237, 437 (1994). HSP60 mtHsp60p Mitochondrial chaperonin 18. H. Hirling, B. R. Henderson, L. C. Kuhn, EMBO J. 13, HSP10 mtHsp10p Mitochondrial chaperonin 453 (1994). SSC1 mtHsp70p Protein import 19. T. L. Megraw, C. B. Chae, J. Biol. Chem. 268, 12758 (1993). a ATP1 -Subunit of F1-ATPase ATP synthesis, protein import 20. J. Blom, M. J. De Mattos, L. A. Grivell, Appl. Environ. Category III Microbiol. 66, 1970 (2000). ACO1 Mitochondrial aconitase Citric acid cycle 21.I.Dilova,C.-Y.Chen,T.Powers,Curr. Biol. 12, 389 (2002). ALD4 Aldehyde dehydrogenase Ethanol metabolism 22. T. Sekito, Z. Liu, J. Thornton, R. A. Butow, Mol. Biol. IDH1 NADþ-dependent isocitrate Citric acid cycle Cell 13, 795 (2002). dehydrogenase, subunit 1 23. J. J. Tate, K. H. Cox, R. Rai, T. G. Cooper, J. Biol. IDP1 NADPþ-dependent isocitrate Oxidative decarboxylation Chem. 277, 20477 (2002). dehydrogenase of isocitrate 24. J.-J. Lawrence, M. Daune, Biochemistry 15, 3301 (1976). 25.M.C.Kennedy,L.Mende-Mueller,G.A.Blondin, KGD1 2-oxoglutarate dehydrogenase, Citric acid cycle H. Beinert, Proc. Natl. Acad. Sci. U.S.A. 89, 11730 E1 component of KGDC (1992). KGD2 2-oxoglutarate dehydrogenase, Citric acid cycle 26. R. D. Klausner, T. A. Rouault, Mol. Biol. Cell 4, 1 (1993). E2 component of KGDC 27. M. D. Williams et al., J. Biol. Chem. 273, 28510 (1998). LPD1 Dihydrolipoamide dehydrogenase, Citric acid cycle, catabolism of 28. L. J. Yan, R. L. Levine, R. S. Sohal, Proc. Natl. Acad. E3 component of PDHC, KGDC, branched-chain amino acids Sci. U.S.A. 94, 11168 (1997). and BCADC 29. J. L. Bradley et al., Hum. Mol. Genet. 9, 275 (2000). LSC1 Succinate-CoA ligase, a subunit Citric acid cycle 30. We thank R. Lill for the generous gift of antibody to aconitase. Supported by NIH grant nos. GM33510 and PDA1 Pyruvate dehydrogenase, Oxidation of pyruvate GM22525 and by Welch Foundation grant no. I-0642. E1 a-subunit of PDHC PDB1 Pyruvate dehydrogenase, Oxidation of pyruvate Supporting Online Material E1 b-subunit of PDHC www.sciencemag.org/cgi/content/full/307/5710/714/ DC1 Category IV Materials and Methods ILV5 Acetohydroxyacid Biosynthesis of Val, Ile, and Leu SOM Text reductoisomerase Figs. S1 to S6 ILV6 Acetolactate synthase Biosynthesis of Val, Ile, and Leu Table S1 regulatory subunit CHA1 L-serine/L-threonine deaminase Catabolism of hydroxy amino acids 14 October 2004; accepted 13 December 2004 10.1126/science.1106391

www.sciencemag.org SCIENCE VOL 307 4 FEBRUARY 2005 717 R EPORTS ship, we performed artificial selection on the FW/BS allometry (20). The FW/BS allometry Natural Selection and evolved rapidly, diverging È2SDineach direction relative to that of the control lineage Developmental Constraints to produce distinct, novel phenotypes EFig. 2; discriminant function analysis correctly in the Evolution of Allometries classified 94.8% of females from generation 13 (–2 log likelihood 0 107.4; N 0 766, W. Anthony Frankino,1*. Bas J. Zwaan,1 David L. Stern,2 replicates pooled)^. The response to selection Paul M. Brakefield1 resulted almost entirely from changes in FW (Fig. 3); BS changed in the appropriate In animals, scaling relationships between appendages and body size exhibit direction in only one lineage (–FW/þBS, 0 0 high interspecific variation but low intraspecific variation. This pattern could lineage E; F1,11 5.55, P 0.038) (20). This result from natural selection for specific allometries or from developmental extreme asymmetry in the contribution of constraints on patterns of differential growth. We performed artificial each trait to the evolution of the allometry selection on the allometry between forewing area and body size in a butterfly was unexpected, as both individual FW and to test for developmental constraints, and then used the resultant increased body size exhibited very similar realized range of phenotypic variation to quantify natural selection on the scaling heritabilities (Fig. 1), indicating adequate and relationship. Our results show that the short-term evolution of allometries is equivalent genetic variation in both traits. not limited by developmental constraints. Instead, scaling relationships are Moreover, the observed pattern of response shaped by strong natural selection. is not due to differences in the phenotypic variance between the traits, because they were Among species, populations, and even sexes, evolve to maintain the relative size of traits subject to similar indirect selection pressures morphological traits exhibit an impressive will then produce developmental constraints in all but one case (Fig. 3). A low frequency diversity of scaling relationships with body Eas defined in (10)^ that must be overcome if of alleles in our starting population that affect size; most traits scale positively with body allometries are to evolve. Here, we present BS independently of FW or a sieving out of size, although the rate at which trait size empirical data addressing the relative roles of key alleles that affect FW but not BS could changes with overall size often differs from natural selection and developmental con- account for the pattern. In any case, the rapid isometry and can even be nonlinear (1, 2). straints in the evolution of the allometry evolution of the allometry demonstrates a This is particularly true of insects, which between forewing area and body size in the surprising absence of developmental con- exhibit extremes in trait–body size allome- butterfly Bicyclus anynana. straints restricting change in this scaling tries (3). This extreme variation among In the context of the evolution of allom- relationship. However, the pattern of response groups is in marked contrast to the extent of etries among morphological traits, the scaling exhibited by FW and BS indicates a strong variation within groups; typically, individu- relationship between wing area and body size als within these groups exhibit low variation (i.e., the ratio of body size to wing area, or around some average allometry, reflecting a Bwing loading[) is of interest, because it is tight scaling between body parts and overall ecologically important and taxonomically di- size Ee.g., (4–7)^. verse (13, 14). The size of the wings and Although these patterns have long been flight musculature relative to body mass af- recognized (2, 8), surprisingly little is fects flight performance directly (14), as well known about the evolution of scaling re- as indirectly through thermoregulatory effects lationships (3, 9); in particular, the relative while basking or during ectotherm flight (13). importance of processes shaping their evo- Lepidoptera have the lowest average wing lution is largely uninvestigated (10). Pre- loading among flying insects (15) and exhibit sumably, tight adherence to particular lineage-specific, seasonal morph–specific, or allometries results from external selection sex-specific scaling relationships associated against traits with atypical or nonfunctional with life historical or behavioral correlates Ee.g., relative sizes. Such selection is predicted to (6, 16–20)^. As with most insects, adult body favor the evolution of genetic and develop- size in B. anynana is a highly plastic trait (21), mental systems that properly scale the and forewing area (FW) exhibits a strong, Fig. 1. Response to artificial selection on absolute growth of traits across body sizes, main- positive phenotypic correlation with total body trait size. Forewing area (FW, triangles indicate di- taining functional size relationships in the mass (BS) across the natural range of body rection of selection) and pupal mass (PM, squares) face of environmental and genetic variation size (Pearson correlation coefficient 0 0.86, are shown relative to control means (horizontal (11, 12). However, this scenario presents a N 0 691 stock population females, P 0 0.0001). line) as a function of the cumulated selected dif- paradox: The proximate mechanisms that Moreover, artificial selection for changes ferential. Regression was used to calculate the realized heritability for each trait. Pupal mass in- in FW and pupal mass revealed a genetic È È 0 creased 1.5 SD and decreased 1SDinsix 1Section of Evolutionary Biology, Institute of Biology, correlation between these traits (r 0.75) generations. Realized heritabilities were moder- Leiden University, P.O. Box 9516, 2300 RA Leiden, (Fig. 1) (20). ate in each direction (þPM h2 0 0.28, –PM h2 0 Netherlands. 2Department of Ecology and Evolution- The strong genetic correlation between FW 0.16). FW responded rapidly and asymmetrically ary Biology, Princeton University, Princeton, NJ and body size should constrain their indepen- to selection, increasing È1.5 SD and decreasing È 08544, USA. dent evolution (22), inhibiting phenotype 0.5 SD relative to controls. Realized heritabil- *Present address: Department of Ecology and Evolu- evolution in a direction perpendicular to that ities were moderate and similar to those for pupal mass [þFW h2 0 0.38, –FW h2 0 0.16; note tionary Biology, Princeton University, Princeton, NJ of the wild-type allometry (23). To determine 08544, USA. that these values are twice the slope of the .To whom correspondence should be addressed. whether such internal constraints limit the regression for FW because only females were E-mail: [email protected] short-term evolution of the scaling relation- selected in these lineages (20)].

718 4 FEBRUARY 2005 VOL 307 SCIENCE www.sciencemag.org R EPORTS bias Eor developmental constraint (10)^ in how males to females (20, 29). In both trials, males prolonged bouts of chasing both other males these traits respond indirectly to direct selec- with the wild-type phenotype acquired three and females, as they do in nature (29), which tion on their scaling relationship. Hence, the times as many matings as did males from both suggests that the lower fitness of treatment allometry itself is not developmentally con- phenotype classes with novel wing loadings males may result from decreased locomotor strained; what does appear to be constrained is (Fig. 4; trial 1, G 0 30.2, P G 0.001; trial 2, G 0 performance (20). Intrasexual competition the way in which the individual components 18.381, P G 0.001). These results demonstrate among male phenotypes may also play a contribute to the evolution of this complex strong stabilizing selection favoring the natural role; þFW/–BS and –FW/þBS males may phenotype. scaling relationship between forewing and be excluded from females in the presence of Our results, together with the few other body size in B. anynana. superiorly flying, wild-type males. Alterna- studies that have used artificial selection to Survival among male phenotype classes tively, wild-type males may be selected by alter scaling relationships between morpho- (recapture rates) did not differ (trial 1, G 0 females because of favored signals produced logical traits in insects (24–26), indicate that 0.110, P 0 0.947; trial 2, G 0 0.641, P 0 during courtship or flight (29) or because even strong genetic correlations do not 0.726), a finding consistent with results from treatment males are less appealing visually. constrain phenotype evolution in the short manipulative studies of wing loading in free- In any case, because treatment and wild-type term. It seems that the developmental basis flying butterflies Ee.g., (15, 30, 31)^. Because males were drawn from the same outbred of these genetic correlations is more impor- survival was the same among male pheno- populations, any female preference must be tant than their strength in determining the types, the higher fitness of wild-type males largely unrelated to male genetic background response to selection (27). In particular, must be due to other, nonexclusive, selective in our experiment. Regardless of the cause of under novel selection regimes such as the factors. In the greenhouse, males engage in the higher fitness of wild-type males, we artificial one we imposed, the developmental 1. 5 program coordinating the growth of the Lineage G Lineage H Fig. 3. Indirect response individual traits may influence how these of mean forewing area 1 traits and the relationship between them (triangles indicate direc- tion of FW selection rel- evolves (28). 0. 5 ative to fresh mass) and The lack of developmental constraints on fresh mass (squares) to the evolution of the allometry motivated us to 0 direct selection on their scaling relationship. Val- determine the pattern of natural selection on -0.5 wing loading. To examine the fitness con- ues are plotted relative to control lineage means sequences of deviating from the wild-type -1 (horizontal dashed line)

FW/BS scaling relationship, we measured the Wing area or fresh mass (STD) -1.5 as a function of the cu- mating success of competing wild-type con- mulative indirect selec- trol and novel-phenotype males (two treat- tion on each trait. The 1. 5 ment male classes, þFW/–BS and –FW/þBS) Lineage E Lineage F average indirect response of each trait is shown by in a spacious, naturally planted, tropical green- 1 house. Treatment and control males taken from the individual regression 0. 5 line for that trait. In reciprocal crosses of the replicated lineages of lineage F, divergence in each selected direction were selected for 0 the values did not occur inclusion in the experiment on the basis of until generation 4; hence, their static allometries (20). Hence, all males -0.5 the mean indirect re- came from similarly outcrossed populations, sponse to selection is and treatment and control males were drawn -1 shown from generation 3 onward for this repli- from the same genetic background. Mating suc- Wing area or fresh mass (STD) -1.5 cate. Cartoons repre- cess was determined by the transfer of phe- 02460246sent the selected target notype class–specific colored powder from Cumulative indirect selection differential (STD) phenotype.

Fig. 2. Scatterplots of static al- lometries of individuals from line- ages selected for changes in FW/BS and the evolution of the mean allometry. (A) Phenotype distribu- tions of lineages selected for changes in forewing–body size scaling. Each selected population is shown as a different symbol; repli- cates of a selection direction have the same shading. The mean allom- etry of each selected direction (replicates combined) is shown as the model II regression through the points and is enclosed by a 95% confidence ellipse. (B) Realized her- itabilities of the mean allometries. Mean phenotype (through which the mean allometry passes) for each lineage is shown relative to control values (horizontal dashed line) as a function of the cumulative selection differential. Mean heritabilities (þFW/–HW 0 0.18; –FW/þHW 0 0.24) are equal to twice the slope of the regressions fit to each selected direction (95% confidence intervals shown by dashed lines). Target phenotypes are represented by cartoons in both panels.

www.sciencemag.org SCIENCE VOL 307 4 FEBRUARY 2005 719 R EPORTS

Function, Evolution (Princeton Univ. Press, Princeton, NJ, 2000). 14. S. Vogel, Comparative Biomechanics (Princeton Univ. Press, Princeton, NJ, 2003). 15. J. G. Kingsolver, Evolution 53, 1479 (1999). 16. For example, low-flight butterflies such as non- dispersers or perching, sit-and-wait males have lower ratios of flight muscle mass to wing size than do stronger flying butterflies such as dispersing individ- uals or males that actively patrol for females [reviewed in (19)]. 17. R. E. Strauss, Evolution 44, 86 (1990). 18. P. Wickman, Evolution 46, 1525 (1992). 19. H. Van Dyck, E. Matthysen, Trends Ecol. Evol. 14,172 (1999). 20. See supporting data on Science Online. 21. R. E. Kooi, P. M. Brakefield, Entomol. Exp. Appl. 80, 149 (1996). 22. D. S. Falconer, T. F. C. MacKay, Introduction to (Addison-Wesley Longman, Essex, UK, ed. 4, 1997). Fig. 4. Distributions of static allometries for FW/BS allometries and relative mating success of three 23. D. Schluter, Evolution 50, 1766 (1996). male phenotype classes. (A) Distributions of individual males with þFW/–BM, wild type, and –FW/þBM 24. K. E. Weber, Genetics 126, 975 (1990). phenotypes included in the experiment (extreme phenotypes are represented by cartoons). Solid circles 25. G. S. Wilkinson, Genet. Res. 62, 213 (1993). denote novel phenotype classes; open circles denote wild-type controls. (B)Matingsuccessofeach 26. D. J. Emlen, Evolution 50, 1219 (1996). phenotype class. Columns indicate percentage of recaptured females that mated with males in each class 27. J. B. Wolf, W. A. Frankino, A. F. Agrawal, E. D. Brodie III, and are shown with 95% confidence intervals based on a bimodal distribution. Numbers in the columns A. J. Moore, Evolution 55, 232 (2001). indicate the number of males recaptured in each group. Data from replicate trials are indicated by similar 28. S. H. , Evolution 52, 647 (1998). 29. M. Joron, P. M. Brakefield, Nature 424, 191 (2003). shading (shared between panels). 30. R. B. Srygley, J. G. Kingsolver, Biol. J. Linn. Soc. 70, 707 (2000). 31. J. G. Kingsolver, R. B. Srygley, Evol. Ecol. Res. 2, 593 2. J. S. Huxley, Problems of Relative Growth (Methuen, have documented strong stabilizing selection (2000). London, 1932). on male wing loading. 32. K. Koops provided essential assistance in rearing 3. D. J. Emlen, H. F. Nijhout, Annu. Rev. Entomol. 45, caterpillars, with N. Wurzer and colleagues providing Our findings indicate that it is not internal 661 (2000). maize plants. C. Allen, P. Beldade, and R. Repasky made 4. D. Burkhardt, I. de la Motte, Entomol. Gen. 12,221 developmental constraints, but rather exter- helpful comments on the manuscript; R. Repasky and (1987). nal natural selection, that is the primary A. Buerkle helped with statistics and computer code. In 5. W. G. Eberhard, E. E. Gutierrez, Evolution 45,18(1991). Leiden, we thank R. de Jong and the National Museum force shaping the short-term evolution of 6. R. E. Strauss, in Ordination in the Study of Morphol- of Natural History for access to specimens and the morphological allometries in insects. How- ogy, Evolution, and Systematics of Insects: Applica- Hortus Botanicus for access to the tropical greenhouse. tions and Quantitative Genetic Rationales,J.T. ever, the surprising bias in the morphological Supported by Leiden University and by a 2001 NSF Sorensen, R. Foottit, Eds. (Elsevier Science, Amster- Bioinformatics Postdoctoral Fellowship (W.A.F.). basis of how the allometry evolved suggests dam, 1992), pp. 157–179. that development may strongly influence 7. D. J. Emlen, Science 291, 1534 (2001). Supporting Online Material how individual traits respond to selection 8. S. J. Gould, Biol. Rev. 41, 587 (1966). www.sciencemag.org/cgi/content/full/307/5710/718/ on their scaling relationships. 9. D. L. Stern, D. J. Emlen, Development 126, 1091 DC1 (1999). Materials and Methods 10. J. Maynard Smith et al., Q. Rev. Biol. 60, 265 (1985). Figs. S1 and S2 11. G. P. Wagner, Am. Zool. 36, 36 (1996). References References and Notes 12. G. P. Wagner, G. Booth, H. Bagheri-Chaichian, Evolution 1. D. W. Thompson, On Growth and Form (Cambridge 51, 329 (1997). 20 September 2004; accepted 11 November 2004 Univ. Press, Cambridge, 1917). 13. R. Dudley, The Biomechanics of Insect Flight: Form, 10.1126/science.1105409

and progenitor cells in aging of multiple organ Mechanisms of Hair Graying: systems has been suggested in mice defective in DNA damage repair and telomere mainte- Incomplete Melanocyte Stem nance (3), but melanocytes may be unique in that the oxidative chemistry of melanin biosynthesis can be cytotoxic (4). This led to Cell Maintenance in the Niche the suggestion that differentiated, pigmented Emi K. Nishimura,1*. Scott R. Granter,2 David E. Fisher1* melanocytes (rather than their unpigmented progenitors) are specifically targeted in hair Hair graying is the most obvious sign of aging in humans, yet its mechanism is graying (5, 6). The recent discovery of largely unknown. Here, we used melanocyte-tagged transgenic mice and aging unpigmented melanocyte stem cells, distinctly human hair follicles to demonstrate that hair graying is caused by defective self- located within the hair follicle (7), creates an maintenance of melanocyte stem cells. This process is accelerated dramatically opportunity to determine whether the process with Bcl2 deficiency, which causes selective of melanocyte stem of hair graying arises specifically from cells, but not of differentiated melanocytes, within the niche at their entry into changes in differentiated melanocytes or the the dormant state. Furthermore, physiologic aging of melanocyte stem cells stem-cell pool that provides them. was associated with ectopic pigmentation or differentiation within the niche, a Stem cells are maintained in the niche process accelerated by mutation of the melanocyte master transcriptional microenvironment (8). Hair follicles contain a regulator Mitf. well-demarcated structure for the stem-cell niche (within the lower permanent portion), Qualitative and quantitative changes in stem although the changes are poorly understood whereas differentiated melanocytes reside in and progenitor cells have been implicated in and the process of stem-cell aging has not the hair bulb (at the base of the transient physiological (chronological) aging (1, 2), been visually observed. Involvement of stem portion of the hair follicle) (Fig. 1A) (7, 9).

720 4 FEBRUARY 2005 VOL 307 SCIENCE www.sciencemag.org R EPORTS Hair follicles are constantly renewing, with Fig. 1. Differentiated alternating phases of growth (anagen), regres- melanocytes are lost in sion (catagen), and rest (telogen) (fig. S1). the hair bulb of Bcl-2 Taking advantage of the spatial segregation of deficient mice. (A)Hair follicle structure. Mela- the stem versus differentiated cell compart- nocyte stem cells (blue ments (7), we used melanocyte-targeted (Dct) dots) are in the lower lacZ transgenic mice (7, 10, 11) to examine permanent portion (light the impact of aging on these melanocyte blue): the bulge (Bg) compartments. area in pelage follicles Among hair graying models, the melano- and the lower enlarge- j/j vit/vit ment (LE) in whisker cyte lineage in Bcl2 (12)andMitf (13) follicles. APM, arrector mice show relatively selective hair graying pili muscle (Figure 1A compared with mouse models of syndromic legend in SOM). (B) premature aging, which affect numerous cell Appearance of Bcl2j/j lineages (14, 15). Hair graying in the Bcl2j/j mouse at P58. (C) Hair background has been suggested to arise by graying of whiskers in Bcl2j/j mouse at chemical cytotoxity of melanin synthesis P39. (D and E) Distri- (5, 12, 16, 17). Distribution and morphology bution of lacZþ cells of melanoblasts among Bcl2j/j, Bcl2–/þ, and (melanocytes) in P39 Bcl2þ/þ mice were normal during early Bcl2–/þ and Bcl2j/j development (fig. S2, a to h). Bcl2j/j mice mice carrying the Dct- gray after the first hair molting (Fig. 1, B and lacZ transgene. Pig- mented melanocytes in C) with white hairs. Histologically, differ- the bulb (Bb) [arrows in entiated melanocytes were almost completely (D)] and lacZþ melano- absent in Bcl2j/j pelage (body hair) or blasts in the Bg [arrow- whisker follicles (Fig. 1, E and G) compared head in (D); the inset with Bcl2–/þ (Fig. 1, D, F, and H) or Bcl2þ/þ shows the magnified (18) follicles at postnatal day 39 (P39). view] are completely lost in Bcl2j/j follicles Albino background did not protect against j j (E). Double arrows indi- melanocyte loss in Bcl2 / mice (Fig. 1I), cate the level of Bb or suggesting that melanin synthesis is un- Bg. Magnification is necessary for this melanocyte disappearance. 100.(F to I)Whole- In addition, Bcl2j/j follicles in the second mount lacZ staining of hair cycle lack both differentiated melano- the bulb of whisker fol- licles from Bcl2–/þ and cytes in the hair bulb and undifferentiated Bcl2j/j þ in black and Dct-lacZ melanoblasts in the stem-cell white (albino: Tyrc-2j/c-2j) niche (located at the bulge area in pelage fol- backgrounds at P40. licles) (Fig. 1, D and E, and fig. S2, i and j), Loss of Dct-lacZþ suggesting that Bcl2 might be important for melanocytes was de- survival of melanocyte stem cells. tected in the bulb of Bcl2j/j whisker fol- Looking earlier at P6.5, when hair follicle j j licles regardless of al- morphogenesis is almost complete, Bcl2 / bino background. follicles appear normal (Fig. 2B). In contrast, Bcl2j/j follicles at P8.5 showed sudden, nearly complete loss of melanoblasts in the (fig. S2, k to n). At this stage, niche melano- stem cells at the time of their transition into niche (bulge area, Fig. 2D), whereas the blasts undergo a morphologic change from the dormant state in the niche and could number of melanocytes in the hair bulb did a dendritic shape into a slender, oval potentially be responsible for certain forms not show significant differences between shape with shrinkage to maximal nuclear/ of human presenile hair graying, although Bcl2–/þ and Bcl2j/j mice (Fig. 2E). In both cytoplasmic ratio upon entry into the dormant no direct supporting evidence has been re- pelage and whisker follicles from Bcl2j/j state (Fig. 2, F and G). This change in mor- ported thus far. animals, disappearance of niche melano- phology was seen cyclically at corresponding In contrast to Bcl2j/j,theMitf vit/vit (13) blasts begins at stage 6 of hair follicle mor- stages of subsequent cycles (18). Apoptosis of graying mouse model exhibited a gradual phogenesis Estandardized hair follicle stages melanocyte stem cells was observed at the decrease of melanocyte stem cells rather than basedon(19)^, and by stage 8 they are gone same stage on the albino or black background abrupt loss (figs. S4 and S6). This strain con- (Tyr c-2j/c-2j) in both pelage and whisker hair tains a mild hypomorphic mutation in Mitf, 1 Department of Pediatric Hematology/Oncology, follicles (Fig. 2, H to M). The same pattern of the melanocyte master transcriptional regula- Melanoma Program in Medical Oncology, Dana-Farber cell loss was detected by using Dct-lacZ, KIT tor E(20, 21) and references therein^.Atearly Cancer Institute, Boston Children’s Hospital, Harvard þ Medical School, Boston, MA 02115, USA. 2Department (c-Kit) (KIT), or microphthalmia-associated to mid-anagen of the third hair cycle, lacZ of , Brigham and Women’s Hospital, transcription factor (MITF) as markers (fig. S3 cells left in the niche of Mitf vit/vit pelage Harvard Medical School, Boston, MA 02115, USA. and Fig. 3, A and B). On the other hand, follicles and Mitf vit/þ whisker follicles often *To whom correspondence should be addressed. melanocytes in the epidermis and dermis produced melanin pigment and exhibited a E-mail: [email protected] (E.K.N.); of hairless skin (e.g., tail and soles) sur- bipolar or dendritic morphology (Fig. 3, C [email protected] (D.E.F.) vived throughout the hair regeneration cycle and D, and fig. S4, j and s). These pigmented .Address after 9 February 2005: Department of Der- matology, Hokkaido University Graduate School of (fig. S2, o and p). These findings indicate cells are unusual because the niche of wild- Medicine, N15, W7, Sapporo 060-8638, Japan. that BCL2 selectively protects melanocyte type controls contains only unpigmented mel-

www.sciencemag.org SCIENCE VOL 307 4 FEBRUARY 2005 721 R EPORTS anocyte stem cells. We provisionally use the and correlated closely with aging (Fig. 3, F significantly with aging, thus serving as a term ectopic pigmentation or differentiation and G). Thus, self-maintenance of melano- control population in these studies (fig. S7). for this reproducibly observed population be- cyte stem cells is essentially complete in Follicles from intermediate-aged individu- cause it is uncertain by which pathway these young animals but becomes defective with als (40 to 60 years old) revealed intermediate cells became pigmented, although they were aging. loss of bulge melanoblasts (Fig. 4, C and J). absent in age-matched controls whose niche We also analyzed the distribution of Bulge melanoblasts were found more in melanoblasts remain undifferentiated (Fig. 3E melanoblasts in aging human hair follicles pigmented follicles than in gray follicles and fig. S4u). with the use of MITF immunostaining (Fig. (18), as shown recently with PMEL17þ bulge Physiologic (senile) aging in mice also 4). MITFþ small unpigmented melanoblasts melanoblasts of middle-aged individuals (27). produces hair graying (fig S5), which could be were found in the outer root sheath prefer- In addition, as with aged or Mitfvit mouse caused by loss of melanocyte stem cells. In- entially around the bulge area where the follicles, ectopically pigmented MITFþ cells deed during physiologic aging, niche melano- arrector pili muscle attaches below the level were occasionally observed in the bulge area blasts (lacZþ) were lost in a gradual and of the sebaceous gland (Fig. 4, A to C), or just below. These cells closely resembled progressive fashion (Fig. 3, F and G). More- similar to previously described amelanotic the dendritic melanocytes described by over, whole-mount cross sections of 8-month- melanocytes (22, 23) that express PMEL17 Narisawa et al. in the bulge area of human old follicles revealed pigment-containing (24, 25), a transcriptional target of MITF follicles (28). The ectopically pigmented or melanocytes within the stem-cell niche in (26). These cells have been suggested to be a differentitated melanocytes were seen exclu- addition to their scattered distribution in the reservoir population for differentiated mela- sively in middle-aged follicles but did not outer root sheath below the niche in whisk- nocytes (23) and exhibit very similar mor- accumulate in the bulge area, suggesting that er follicles (Fig. 3, H and I, and fig. S5, k to phology to melanocyte stem cells in mice. they are not self-maintaining. n). The appearance of these pigmented Whereas MITFþ immature melanoblasts Our results demonstrate that Bcl2 is melanocytes in the niche is reminiscent of were abundant in follicles from 20- to 30- selectively critical for maintenance of mela- pigmented niche melanocytes observed year-old subjects (2 to 3% of the total basal nocyte stem cells, specifically for entry into during the accelerated graying of Mitf-vit keratinocytes in the bulge area), they were the dormant state. Bcl2 was previously mutants. Quantitative analysis revealed that absent from most hair follicles of 70- to 90- shown to modulate hematopoietic stem-cell the presence of these cells was accompanied year-old subjects (Fig. 4J). MITFþ melano- pool size (29). Different lineages might use by simultaneous loss of the typical unpig- cytes in the uppermost area (infundibulum) distinct antiapoptotic mechanisms to resist mented Dct-lacZþ melanoblasts in the niche of the outer root sheath did not decrease the specific stress signals for dormancy.

Fig. 2. Loss of Bcl2j/j melanocyte stem cells upon entry into the Terminal deoxynucleotidyl transferase–mediated deoxyuridine tri- dormant state. (A to D) Distribution of Dct-lacZþ melanoblasts (arrows) phosphate nick end labeling (TUNEL), lacZ, and 4¶,6¶-diamidino-2- in the Bg (top double arrow) of pelage follicles at P6.5 and P8.5. Whereas phenylindole (DAPI) staining of stage 7 skin from P6.5 Bcl2j/j (H) and Bb melanocytes appear largely unchanged (bottom double arrow), bulge P6.5 Bcl2þ/j (I) mice. Arrowheads show apoptotic inner root sheath melanoblasts are lost in Bcl2j/j follicles at P8.5 [compare (D) with (C)] keratinocytes. The inset area, marked with the arrow in (H), shows an but not at P6.5 [compare (B) with (A)]. (E) Comparison of the total apoptotic melanoblast. The top inset on the right shows the merged view number per field of Dct-lacZþ melanoblasts in the bulb versus in the bulge for TUNEL (green) and DAPI (blue). The bottom inset shows the merged plus subbulge of Bcl2j/j and Bcl2j/þ pelage follicles at P8.5 on 7-mm view for TUNEL (green) and LacZ (red). Distribution of Dct-lacZþ sections (magnified 100). KIT expression matches Dct-lacZþ in bulge melanoblasts in the niche: Bg of pelage hair follicles (J and K) and LE melanoblasts [stage 6 (F)andstage8(G)] of Bcl2þ/þ animals (double arrow) of whisker hair follicles [(L and M), double arrows] from (magnification, 630). Cell size is diminished from stage 6 to stage 8. P8.0 mice with white backgrounds (Tyr c-2j/c-2j).

722 4 FEBRUARY 2005 VOL 307 SCIENCE www.sciencemag.org R EPORTS Fig. 3. Effect of aging Although melanin biosynthesis has been ele- and Mitf mutation on gantly shown to be cytotoxic in the context melanocyte stem cells. of a certain genetic mutation (6), stem-cell (A and B)Coincident expression of Dct-lacZ, disappearance in Bcl2 null mice does not KIT, and MITF in hair require melanogenesis. Bcl2 is a transcrip- follicle melanoblasts or tional target of MITF (30), but Bcl2 does not melanocytes (magnifi- appear to fully account for melanocyte loss cation, 200). (See fig. in the context of the weakly hypomorphic S3 for more details.) (C) Mitf vit/vit allele. Ectopically pigmented melanoblasts (lacZþ, Our data suggest a previously unknown blue) in the bulge region pathophysiologic explanation for hair graying. (arrow) of 3.5-month- Loss of melanocyte stem cells can be observed old Mitf vit/vit follicles. and temporally precedes the loss of dif- (D) Magnified view of ferentiated melanocytes in the hair matrix. pigmented bulge mela- Thus, incomplete maintenance of melanocyte noblasts. (E)Absenceof pigment in lacZþ bulge stem cells appears to cause physiologic hair melanoblasts of age- graying through loss of the differentiated matched Mitfþ/þ fol- progeny with aging. This is associated with licles. (F) Quantitation ectopic melanocyte pigmentation or differ- of niche melanocytes entiation within the niche. Possible expla- þ (lacZ ), either un- nations include premature differentiation or pigmented (classical activation of a senescence program Ewhich stem cells, blue) or ^ ectopically pigmented induces pigmentation in vitro (31) .Ac- (green), in LE of whisker celeration of this process in Mitfvit follicles follicles (positions a2 implicates MITF in the self-renewal of and a3) (see fig. S1i for melanocyte stem cells. The precise roles for positions). (G)Number stem-cell apoptosis versus ectopic differentia- of unpigmented niche melanoblasts in whisker tionremaintobedeterminedbutmaysimilarly follicles (positions a2, contribute to stem-cell loss in other aging or- a3, c5, and d5) with black, gray, and white hair in 18- to 22-month-old (18–22M) wild-type mice. gan systems. Asterisk indicates statistical significance (P G 0.01). (H and I) Ectopically pigmented melanoblasts in the niche (LE of whisker follicles) of aging wild-type mice (whole-mount view). References and Notes 1. M. A. Sussman, P. Anversa, Annu. Rev. Physiol. 66,29 Fig. 4. Melanoblast and (2004). 2. G. Van Zant, Y. Liang, Exp. Hematol. 31, 659 (2003). melanocyte distribu- 3. K. K. Wong et al., Nature 421, 643 (2003). tion in human hair fol- 4. K. Urabe et al., Biochim. Biophys. Acta 1221,272 licles from different age (1994). groups. Human scalp 5. J. P. Ortonne, J. J. Nordlund, in The Pigmentary System, specimens were immu- J. J. Nordlund, R. Boissy, V. J. Hearing, R. King, J. P. nostainedwithanti- Ortonne, Eds. (Oxford Univ. Press, New York, 1998), bodies against MITF. (A pp. 489–502. and B)MITFþ cells (ar- 6. R. Johnson, I. J. Jackson, Nature Genet. 1, 226 (1992). 7. E. K. Nishimura et al., Nature 416, 854 (2002). rows) are distributed on 8. F. M. Watt, B. L. M. Hogan, Science 287, 1427 (2000). the outer root sheath in 9. E. Fuchs, B. J. Merrill, C. Jamora, R. DasGupta, Dev. the bulge of follicles Cell 1, 13 (2001). from 20- to 30-year- 10. M. A. Mackenzie, S. A. Jordan, P. S. Budd, I. J. Jackson, old individuals. Magnifi- Dev. Biol. 192, 99 (1997). cation in (A), 200;(B), 11. K. R. Fitch et al., Genes Dev. 17, 214 (2003). 630.(C and D)Rep- 12. D. J. Veis, C. M. Sorenson, J. R. Shutter, S. J. Korsmeyer, resentative views of the Cell 75, 229 (1993). 13. A. B. Lerner et al., J. Invest. Dermatol. 87, 299 (1986). bulge from follicles of 14. K. L. Rudolph et al., Cell 96, 701 (1999). 40- to 60- and 70- to 15. J. de Boer et al., Science 296, 1276 (2002); published 90-year-old people, online 11 April 2002 (10.1126/science.1070174). respectively (magnifica- 16. K. Yamamura et al., Cancer Res. 56, 3546 (1996). tion, 630). (E)Sche- 17. D. J. Tobin, R. Paus, Exp. Gerontol. 36, 29 (2001). matic for human hair 18. E. K. Nishimura, S. R. Granter, D. E. Fisher, unpublished follicle with pigmented data. hair. Immature MITFlow 19. R. Paus et al., J. Invest. Dermatol. 113, 523 (1999). 20. E. Steingrimsson, N. G. Copeland, N. A. Jenkins, Annu. melanoblasts (blue) are Rev. Genet. 38, 365 (2004). located in the lower 21. E. R. Price, D. E. Fisher, Neuron 30, 15 (2001). permanent portion (the 22. W. Montagna, H. B. Chase, Am. J. Anat. 99, 415 (1956). bulge). MITFhigh mela- 23. R. G. Staricco, Ann. N. Y. Acad. Sci. 100, 239 (1963). nocytes are located in 24. T. Horikawa et al., J. Invest. Dermatol. 106,28 epidermis, infundibulum (1996). (If) (brown), and hair 25. S. Commo, B. A. Bernard, Pigment Cell Res. 13, 253 matrix (M, green). ORS, outer root sheath. See Fig. 1A for abbreviations. (F to H) The bulb region of (2000). 26. J. Du et al., Am. J. Pathol. 163, 333 (2003). follicles from different age groups. Mature melanocytes in the hair matrix express MITF (arrowheads). yo, þ 27. S. Commo, O. Gaillard, B. A. Bernard, Br. J. Dermatol. years old. (I)AnMITF melanocyte that contains abundant melanin granules and long dendrites is 150, 435 (2004). detected in the bulge and subbulge of follicles specifically from middle-aged individuals. (J) The frequency 28. Y. Narisawa, H. Kohda, T. Tanaka, Acta Derm. of MITFþ cells per basal keratinocytes in the bulge. Asterisks indicate statistical significance (P G 0.01). Venereol. 77,97(1997).

www.sciencemag.org SCIENCE VOL 307 4 FEBRUARY 2005 723 R EPORTS

29. J. Domen, S. H. Cheshier, I. L. Weissman, J. Exp. Med. Rowitch for sharing microscopes. E.K.N. wishes Supporting Online Material 191, 253 (2000). to thank Y. Miyachi for continuous encourage- www.sciencemag.org/cgi/content/full/1099593/DC1 30. G. G. McGill et al., Cell 109, 707 (2002). ment and support. E.K.N. was supported by the Materials and Methods 31. E. E. Medrano et al., Mol. Biol. Cell 5, 497 (1994). Shiseido Award in 2002 and the Charles A. King Figs. S1 to S7 32. We thank V. Igras and C. Quigley for technical Trust, Fleet National Bank, a Bank of America Com- assistance; L. Lamoreux for supplying Mitf vit pany, co-trustee (Boston, MA). Supported by grant 26 April 2004; accepted 8 December 2004 mice;A.Miller,C.Hershey,andY.Choiforcriti- no. AR43369 from NIH to D.E.F. D.E.F. is the Nirenberg Published online 23 December 2004; cal reading of the manuscript; members of the in pediatric oncology at Dana-Farber Can- 10.1126/science.1099593 Fisher laboratory for helpful discussions; and D. cer Institute. Include this information when citing this paper.

tional modules (9), we found that all but two Dynamic Complex Formation of them were identified by our approach (bet- ter than random at P G 10–30). The only ex- During the Yeast Cell Cycle ceptions were the anaphase-promoting complex (APC), which can only be detected with a Ulrik de Lichtenberg,1* Lars Juhl Jensen,2* less stringent interaction cutoff, and the Skp1p/ Søren Brunak,1 Peer Bork2,3. Cullin/F-box protein complex (SCF), which appears to be the only cell cycle–related pro- To analyze the dynamics of protein complexes during the yeast cell cycle, we tein complex without a periodically expressed integrated data on protein interactions and gene expression. The resulting subunit. For completeness, these two com- time-dependent interaction network places both periodically and constitu- plexes were added to the network. Our extrac- tively expressed proteins in a temporal cell cycle context, thereby revealing tion procedure produces comparable results previously unknown components and modules. We discovered that most com- even if the curated MIPS complexes are plexes consist of both periodically and constitutively expressed subunits, which excluded entirely from the analysis or if the suggests that the former control complex activity by a mechanism of just- specific extraction criteria are changed, show- in-time assembly. Consistent with this, we show that additional regulation ing that the method is robust and has much through targeted degradation and phosphorylation by Cdc28p (Cdk1) specif- higher coverage than methods of comparable ically affects the periodically expressed proteins. accuracy (13). The derived cell cycle network (Fig. 1, in- Most research on biological networks has been were selected (13). These were further side circle) contains 29 heavily intraconnect- focused on static topological properties (1), filtered with information on subcellular ed modules; that is, complexes or groups of describing networks as collections of nodes localization (14) to exclude interactions complex variants that exist at different time and edges rather than as dynamic structural between proteins annotated to incompatible points during the yeast cell cycle. In addition entities. Here we focus on the temporal as- compartments (13); no curated MIPS inter- to rediscovering many known cell cycle mod- pects of networks, which allows us to study the actions were lost because of this filtering. ules, our approach enables us to place more dynamics of protein complex assembly during The topology-based scoring scheme, filter- than 30 poorly characterized proteins in the the Saccharomyces cerevisiae cell cycle. ing, and extraction criteria reduced the error cell cycle network and to predict new unex- Our integrative approach combines protein- rate for interactions by an order of magni- pected cell cycle contexts for other proteins protein interactions with information on the tude to only 3 to 5% (13). (13). The network contains 31 isolated binary timing of the transcription of specific genes In the extracted network (Fig. 1), we in- complexes, many of which involve proteins during the cell cycle, obtained from DNA cluded, in addition to the periodically ex- of unknown function, such as Yml119p and microarray time series (2, 3). From the latter, pressed (Bdynamic[) proteins, constitutively Yll032p, which interact and are both putative we derived a quality-controlled set of 600 expressed (Bstatic[) proteins that preferen- Cdc28p substrates (16) expressed close in

periodically expressed genes, each assigned tially interact with dynamic ones (13). The time in G2 phase (13). to the point in the cell cycle where its resulting network consists of 300 proteins As an example of the value of combining expression peaks (4). We then constructed a (Fig. 1, inside circle), including 184 dynamic temporal data with protein-protein interactions, physical interaction network for the correspond- proteins (colored according to their time of the network reveals a binary complex consist- ing proteins from yeast two-hybrid screens peak expression) and 116 static proteins (de- ing of the uncharacterized proteins Ymr295p (5, 6), complex pull-downs (7, 8), and cu- picted in white). For 412 of the 600 dynamic and Ydr348p. Because only Ydr295p is dy- rated complexes from the Munich Information proteins identified in the microarray analysis, namic, the static protein Ydr348p can only Center for Protein Sequences (MIPS) database no physical interactions of sufficient reli- be identified as a cell cycle–relevant protein (9). To reduce the error rate of 30 to 50% ability could be found (Fig. 1, outside circle). and placed temporally through the integra- expected in most current large-scale interac- Some may be missed subunits of stable com- tion of the two complementary data types. tion screens (10, 11), all physical interaction plexes already in the network; the majority, Indeed, Ydr348p is a putative Cdc28p target data were combined, a topology-based con- however, probably participate in transient in- (16), and the interaction is further supported by fidence score was assigned to each individ- teractions, which are often not detected by the observation that both proteins localize to ual interaction Eas in the STRING database current interaction assays (15). the bud neck (14). Virtually all complexes con- (12)^, and only high-confidence interactions Although our procedure for extracting in- tain both dynamic and static subunits (Fig. 1), teractions might miss some cellular processes the latter accounting for about half of the 1Center for Biological Sequence Analysis, Technical that are dominated by transient interactions, direct interaction partners of periodically regu- University of Denmark, DK-2800 Lyngby, Denmark. most of the stable complexes should have lated proteins through all phases of the cell 2European Molecular Biology Laboratory, D-69117 3 been captured at least partially. Tandem af- cycle (Fig. 2). Transcriptional regulation thus Heidelberg, Germany. Max-Delbru¨ck-Centre for Mo- lecular Medicine, D-13092 Berlin, Germany. finity purifications alone should identify at influences almost all cell cycle complexes and least half of the subunits for 87% of the thereby, indirectly, their static subunits. This *These authors contributed equally to this work. .To whom correspondence should be addressed. known yeast complexes (7). Compared with implies that many cell cycle proteins cannot E-mail: [email protected] the known cell cycle complexes and func- be identified through the analysis of any sin-

724 4 FEBRUARY 2005 VOL 307 SCIENCE www.sciencemag.org R EPORTS

M/G1

M G1

Regulation MCM/ORC Protein of kinase A

DNA replication Glycogen synthesis

Nucleosome/ APC Mitotic exit bud formation DNA polymerase

Cdc28-cyclin

Histones Cation transport DNA replication SCF Pho85- and repair cyclin

Sister chromatid cohesion

Tubulin Septin related filaments

Transcription factors SPB Cell wall

G2 S

Fig. 1. Temporal protein interaction network of the yeast mitotic cell without interactions are both positioned and colored according to their cycle. Cell cycle proteins that are part of complexes or other physical peak time and thus also serve as a legend for the color scheme in the interactions are shown within the circle. For the dynamic proteins, the network. More detailed versions of this figure (including all protein time of peak expression is shown by the node color; static proteins are names) and the underlying data are available online at www.cbs.dtu.dk/ represented by white nodes. Outside the circle, the dynamic proteins cellcycle. gle type of experimental data but only through ules. Most notably, the network reveals a Transcription of cell cycle–regulated genes integrative analysis of several data types. module that includes two poorly character- is generally thought to be turned on when or In addition to suggesting functions for in- ized proteins (Nis1p and Yol070p) and links just before their protein products are needed: dividual proteins, the network (Fig. 1) indicates processes related to the nucleosomes with often referred to as just-in-time synthesis. Con- the existence of entire previously unknown mod- mitotic events in the bud (Fig. 3A) (13). trary to the cell cycle in bacteria (17), how-

www.sciencemag.org SCIENCE VOL 307 4 FEBRUARY 2005 725 R EPORTS ever, just-in-time synthesis of entire complexes and numerous smaller complexes or modules supported by binding of the transcription fac- is rarely observed in the network. The only (13). We find that the time map- tors Fkh2p, Mcm1, and Ndd1p, which coop-

large complex to be synthesized in its entirety pings visualized in Fig. 1 are in close agree- eratively activate transcription at the G2/M just in time is the nucleosome, all subunits ment with previous studies on the dynamic transition, exactly when the expression of of which are expressed in S phase to produce formation of individual protein complexes, Mmr1p peaks (21, 22). nucleosomes during DNA replication. suggesting that the timing of transcription of Furthermore, we used the network model Instead, the general design principle ap- dynamic proteins is indicative of the timing of to determine whether the transcriptionally pears to be that only some subunits of each assembly and action of the complexes and regulated subunits are also regulated by phos- complex are transcriptionally regulated in or- modules (Fig. 3) (13). Just-in-time assembly phorylation. The 332 putative Cdc28p tar- der to control the timing of final assembly. would have an advantage over just-in-time gets identified in a recent screen (16) are Several examples of this just-in-time assembly synthesis of entire complexes in that only a indeed much more frequent among the dy- (rather than just-in-time synthesis) are sug- few components need to be tightly regulated namic than among the static proteins in the gested by the network, including the prerepli- in order to control the timing of final com- network Esignificant at P G 10–4 (13)^. In the cation complex (Fig. 3B), complexes involved plex assembly. This would explain the recent cell cycle network (Fig. 1), 27% of the period- in DNA replication and repair, the spindle observation that the periodic transcription of ically transcribed proteins are putative Cdc28p pole body, proteins related to the cytoskeleton, specific cell cycle genes is poorly conserved targets, in comparison to only 8% of the static through evolution (18). For the prereplica- proteins and 6% of all yeast proteins. This tion complex, exactly this variation between link between transcriptional and posttrans- organisms has been shown, although the sub- lational control probably reflects multiple roles: units and the order in which they assemble It allows the control of complex assembly by are conserved (19). dynamic proteins to be fine-tuned, provides Dynamic complex assembly also func- new nonmodified Cdc28p targets in each cell tions as a mechanism for temporal regulation cycle, and regulates the degradation of the of substrate specificity, best exemplified by dynamic proteins through phosphorylation. the association of the cyclin-dependent ki- The latter is supported by an overrepresen- nase Cdc28p with its various transcription- tation of predicted PEST (regions rich in Pro, ally regulated cyclins and inhibitors (Fig. 3C). Glu, Ser, and Thr) degradation signals among Our approach accurately reproduces this key both the dynamic proteins and the putative regulatory system and its temporal dynamics, Cdc28p targets in the network Esignificant at correctly placing each of the Cdc28p inter- P G 10–2 and P G 10–3, respectively (13)^. Cell actions_ partners at their time of function and cycle–regulated transcription thus supplies the capturing even very transient interactions such cell with nonmodified complex components at as phosphorylation and ubiquitination (Fig. 3C). the time of complex assembly, many of which Fig. 2. Interactions during the cell cycle. The Transient interactions are also captured be- are later phosphorylated and specifically tar- number of interactions of dynamic proteins with tween cyclins of the Pho85p system and two geted for degradation. other dynamic proteins (dynamic-dynamic) and M-phase–specific proteins, Swi5p and Mmr1p. With the emergence of new large-scale with static proteins (dynamic-static) is shown as Swi5p has already been identified as a Pho85p- data sets, including assays focused on protein- a function of cell cycle progression. Zero time Pcl2p substrate (20); from the network context, DNA interactions and transient protein-protein corresponds to the time of cell division. The we propose that the uncharacterized phospho- interactions, we expect more of the dynamic large number of interactions during G1 phase reflects a general overrepresentation of genes protein Mmr1p is a target of the Pho85p-Pcl7p proteins to be included in the network. Also, expressed in this part of the cell cycle (Fig. 1). kinase (13). The cell cycle role of Mmr1p is we currently lack information about the life-

Fig. 3. Dynamic modules. Each panel shows a specific module from the are bound to origins of replication throughout the entire division cycle. A

network in Fig. 1, using the corresponding colors of individual proteins. subcomplex of six Mcm proteins is recruited to the ORC complex in G1 phase by (A) Previously unknown module connecting processes related to chromo- a Cdc6p-dependent mechanism; we see the corresponding genes (except MCM6) some structure with mitotic events in the bud. The nucleosome assembly transcribed just before that. Final recruitment of the replication machinery is protein Nap1p is known to shuttle between the nucleus and cytosol and dependent on Cdc45p, which we find expressed in early S phase. More details

regulates the activity of Gin4p, one of two G1-expressed budding-related and case stories are available (13). (C) Cdc28p module, with the different kinases in the module. Nap1p and the histone variant Htz1p connect the cyclins and interactors placed at their time of synthesis. At the end of module to the nucleosome and sister chromatid modules, respectively. Two , the cyclins are ubiquitinated and targeted for destruction by APC poorly characterized proteins, Nis1p and the putative Cdc28p substrate and SCF, reflected in the network by the interaction between Cdh1p and Yol070p (16), are both expressed in mitosis and localize to the bud neck Clb2p. The latter also interacts with Swe1p, which inhibits entry into mitosis (13). (B) Schematic representation of the dynamic assembly of the by phosphorylating Cdc28p in complex with Clb-type cyclins. We also discov- prereplication complex. It contains six static proteins (Orc1p to Orc6p) that ered an uncharacterized protein, Ypl014p, in this well-studied module (13).

726 4 FEBRUARY 2005 VOL 307 SCIENCE www.sciencemag.org R EPORTS time of the observed complexes and modules. 5. T. Ito et al., Proc. Natl. Acad. Sci. U.S.A. 97, 1143 20. V. Measday, H. McBride, J. Moffat, D. Stillman, B. With reliable time series of protein abun- (2000). Andrews, Mol. Microbiol. 35, 825 (2000). 6. P. Uetz et al., Nature 403, 623 (2000). 21. I. Simon et al., Cell 106, 697 (2001). dances, preferably in individual compartments, 7. A.-C. Gavin et al., Nature 415, 141 (2002). 22. T. I. Lee et al., Science 298, 799 (2002). the resolution of this temporal network can 8. Y. Ho et al., Nature 415, 180 (2002). 23. We thank S. Hooper for assistance and M. Knop and be increased considerably, because even in- 9. H. W. Mewes et al., Nucleic Acids Res. 32, 5539 (2004). members of the Bork group for insightful comments 10. A. M. Edwards et al., Trends Genet. 18, 529 (2002). on the manuscript. Supported by grants from the dividual interactions over time could then 11. C. von Mering et al., Nature 417, 399 (2002). Danish National Research Foundation, the Danish be monitored. Moreover, the integrative ap- 12. C. von Mering et al., Nucleic Acids Res. 31,258 Technical Research Council, Novo Nordisk A/S, and proach presented here should be applicable (2003). the European Union. to any biological system for which both in- 13. See supporting information on Science Online for Supporting Online Material details. teraction data and time series are available. www.sciencemag.org/cgi/content/full/307/5710/724/ 14. K. R. Christie et al., Nucleic Acids Res. 32, D311 (2004). DC1 References and Notes 15. L. J. Jensen, P. Bork, Drug Discovery Today: TARGETS Materials and Methods 1. A. L. Barabasi, Z. N. Oltvai, Nature Rev. Genet. 5, 101 3, 51 (2004). Fig. S1 to S3 (2004). 16. J. A. Ubersax et al., Nature 425, 859 (2003). Table S1 2. R. J. Cho et al., Mol. Cell 2, 65 (1998). 17.H.H.McAdams,L.Shapiro,Science 301, 1874 References 3. P. T. Spellman et al., Mol. Biol. Cell 9, 3273 (1998). (2003). 4. U. de Lichtenberg et al., Bioinformatics doi:10.1093/ 18. G. Rustici et al., Nature Genet. 36, 809 (2004). 10 September 2004; accepted 9 December 2004 bioinformatics/bti093 (2004). 19. M. L. DePamphilis, Gene 310, 1 (2003). 10.1126/science.1105103

WT.Furthermore,whenBHKcellsexpressing Escape of Intracellular Shigella GFP-LC3, an autophagosome-specific marker (13, 14), were infected with DicsB or WT, from Autophagy È40% of DicsB was associated with LC3 1 3,6 1,5 signal; bacterial shape was also indistinct Michinaga Ogawa, Tamotsu Yoshimori, Toshihiko Suzuki, compared with WT (fig. S1D). To further 2 4,5 1,6 Hiroshi Sagara, Noboru Mizushima, Chihiro Sasakawa * characterize the DicsB defect, we exploited MDCK cells (epithelial cells from dog kidney) The degradation of undesirable cellular components or organelles, including expressing GFP-LC3 (MDCK/pGFP-LC3 invading microbes, by autophagy is crucial for cell survival. Here, Shigella,an cells), which made it feasible to visualize invasive bacteria, was found to be able to escape autophagy by secreting IcsB cytoplasmic organelles and bacteria (Fig. 1A). by means of the type III system. Mutant bacteria lacking IcsB were The number of LC3-positive DicsB was greater trapped by autophagy during multiplication within the host cells. IcsB did not than that of WT throughout the 1 to 6 hours directly inhibit autophagy. Rather, Shigella VirG, a protein required for after infection. The LC3-positive population of intracellular actin-based motility, induced autophagy by binding to the DicsB had increased 50% by 6 hours, whereas autophagy protein, Atg5. In nonmutant Shigella, this binding is competitively that of WT remained at 10 to 15% (Fig. 1B). inhibited by IcsB binding to VirG. Two hours after infection, WT and DicsB had similar numbers of actin tails. After 4 hours, During the multiplication of microbes within polymerization. Cytoplasmic pathogens may however, the population was decreased in host cells, bacteria become sequestered in thus circumvent autophagic events. DicsB (fig.S2),presumablybecauseDicsB membrane-bound organelles such as phago- IcsB, one of the Shigella flexneri effectors, was within autophagosomes. The LC3-positive somes (1–3). This event is a key component of is secreted by means of the type III secretion population of the DicsB/pIcsB was decreased: host defense against invading microbes. system (TTSS) of cytoplasmic bacteria and it fell to a level as low as that of WT (Fig. 1B). Nevertheless, some invasive bacteria such as located on the bacterial surface (12). The icsB Autophagic events can be triggered by amino Legionella, Salmonella, Mycobacteria,and mutant is fully invasive and able to escape acid starvation (13). MDCK/pGFP-LC3 cells Brucella can block or alter the maturation of from the vacuole but is defective in spreading were infected with DicsB or WT, under amino the phagosome and can reside in vacuoles within host cells (12). acid–starved conditions. LC3-positive bacteria (2–7). Some others such as Shigella (8, 9), To clarify the role of IcsB in promoting in MDCK cells were significantly increased Listeria monocytogenes (10), and Rickettsia infection, we investigated the intracellular from 10 to 16% (WT) and from 23 to 36% conorii (11) can escape from phagosomes behaviors of the icsB mutant (DicsB), (DicsB) in response to amino acid deprivation into the cytoplasm, multiply, and dissemi- YSH6000 (wild type; WT), and DicsB/pIcsB (fig. S3). Conversely, when MDCK cells were nate into neighboring cells by eliciting actin (the icsB complement strain). In baby hamster treated with known inhibitors of autophagy or kidney (BHK) cells, although mutants lacking of lysosomes, such as Wortmannin, 3-methyl- IcsB multiplied as normal for about 3 hours, adenine (3-MA) or bafilomycin-A1 (Baf-A1), 1Department of Microbiology and , 2De- their growth plateaued 4 hours after invasion the LC3-positive DicsB population was mark- partment of Fine Morphology, Institute of Medical (fig. S1A). To characterize intracellular bacte- edly decreased (Fig. 1, C and D). In the Science, University of Tokyo, 4-6-1, Shirokanedai, ria, we introduced green fluorescent protein presence of Baf-A1, fusion of lysosomes with 3 Minato-ku, Tokyo 108-8639, Japan. Department of plasmid (pGFP) into DicsB and WT then autophagosomes containing DicsB was Cell Genetics, National Institute of Genetics, 1111, Yata, Mishima, Shizuoka 411-8540, Japan. 4Department investigated BHK cells infected with bacteria 4 blocked, which would have allowed the of Bioregulation and Metabolism, Tokyo Metropolitan hours after infection. DicsB/pGFP colocalized bacteria to escape into the cytosol. Consist- Institute of Medical Science, 3-18-22, Hon-komagome, with markers for acidic lysosomes (Lysotracker) ently, despite the smaller diameter (G0.15 mm) 5 Bunkyo-ku, Tokyo 113-8613, Japan. Precursory or autophagosomes Emonodancyl-cadaverin of plaques formed by DicsB 2 days after Research for Embryonic Science and Technology 6 (MDC)^, where the bacterial morphology was infection than that of plaques formed by WT (PRESTO), Core Research for Evolutional Science È and Technology (CREST), Japan Science and Technol- indistinct. (fig. S1, B and C). WT cells, on the ( 0.5 mm), the plaque-forming capacity of ogy Agency, Kawaguchi 332-0012, Japan. whole, did not colocalize with the same DicsB was restored by treatment with Baf-A1 *To whom correspondence should be addressed. markers: 37.2% of DicsB bacteria colocalized (fig. S4). Another investigation was made in E-mail: [email protected] with lysosomes compared with only 10.2% of atg5-knockout mouse embryonic fibroblasts

www.sciencemag.org SCIENCE VOL 307 4 FEBRUARY 2005 727 R EPORTS

j/j A B 60 (atg5 MEFs), which are defective in WT j/j icsB WT * autophagy. When atg5 MEFs or normal icsB þ þ icsB/pIcsB MEF cells (atg5 / MEFs) expressing GFP- 50 LC3 were infected with DicsB, even though * LC3-positive DicsB is detectable in normal h LC-3 (%) 40 MEFs, signals were barely detected in the

wit *

d atg5-knockout MEFs (Fig. 1, E and F). te

a 30 Consistently, intracellular growth of DicsB ci o was recovered to the level of WT in atg5j/j ss 20 MEF cells (Fig. 1G), which provided further a evidence that the defective intracellular cteri phenotype of DicsB was associated with

Ba 10 autophagy. MDCK cells infected with DicsB or WT 0 C 1 2 4 6 for 4 hours were examined by thin-section icsB +Wortmannin Time after infection (h) electron microscopy (EM). DicsB was fre- D 40 quently enclosed by lamellar membranous ** WT icsB structures, in striking contrast to the phagocytic 35 membrane surrounding an invading bacterium (Fig. 2A). Occasionally, some bacteria 30 enclosed by lamellar membranes were also h LC-3 (%) 25 surrounded by structures like onion skin (Fig. wit

d 2A, arrows). Six hours after invasion, some te +3-MA +Baf-A1 a 20 bacteria enclosed by an onion skin–like ci o * structure had become indistinct (fig. S5A). ss 15 At the same stage of infection, WT bacteria a 10 generally lacked the lamellar membranes. cteri Instead, most had long actin tails, as seen in Ba 5 motile bacteria (fig. S5B). To confirm that the

0 lamellar membranes surrounding DicsB were untreated 3-MA Wortmannin Baf-A1 autophagic membranes, MDCK/pGFP-LC3 40 cells infected with DicsB 4 hours after E F MEF atg5 +/+ * +/+ -/- infection were analyzed by immunogold EM MEF atg5 MEF atg5 35 MEF atg5 -/- ) with antibody against GFP. The lamellar % ( 30 membrane around DicsB in MDCK/pGFP- s l l LC3 cells was specifically labeled with e c 25 immunogold (Fig. 2B, arrows). Onion skin– e v i t

i like membranous structures associated with

s 20 o the bacterium were also highly labeled with p

3 15 immunogold (Fig. 2B, arrowheads). Thus C L

- intracellular Shigella lacking IcsB readily P

F 10 * succumbs to autophagy within epithelial G cells. 5 To explore the role of IcsB, we assessed whether IcsB is linked to autophagic pro- Fig. 1. Shigella icsB mutant trapped by autophagy. 0 (A) MDCK/pGFP-LC3 cells infected with WT or WT icsB teins. We constructed COS-7 transfectants DicsB for 4 hours were stained with antibody expressing GFP-Atg5, GFP-Beclin (Atg6), atg5 +/+ WT G 35 against Shigella (blue) and rhodamine phalloidin atg5 +/+ icsB GFP-LC3 or Myc in complex with the (red) and (B)thepercentagesofGFP-LC3– atg5 -/- WT vacuolar protein-sorting protein Vps34, that G -/- positive bacteria in (A) were quantified. *P 30 atg5 icsB is Myc-Vps34 (type III phosphatidylinositol 0.0001. (C) In the presence of autophagy 3-kinase), and each cell lysate was placed inhibitors, MDCK/pGFP-LC3 cells infected with x 25 e * DicsB for 4 hours were stained with antibody d in a pull-down assay with glutathione S- n against Shigella (red). In (D), cells described in i transferase (GST) in complex with IcsB. y 20 * t i

(C) are compared with cells infected with WT c i However, under these experimental con- l p

under the same conditions to determine the i

t 15 ditions, none was precipitated by GST-IcsB. G l percentages of GFP-LC3–positive bacteria. *P u Intriguingly, we noted that the LC3 and Atg5 0.0001. (E) atg5-knockout MEFs or normal MEFs M 10 expressing GFP-LC3 infected with DicsB for 4 signals in the vicinity of the bacterial body hours were stained with antibody against Shi- of DicsB tended to be distributed asym- gella (red) and compared with those infected 5 metrically, with signals occasionally accu- with WT. (F) Under the same conditions, they mulating at one pole of the bacterium. were stained with antibody against Shigella. The 0 0 50 100 150 200 250 300 350 Furthermore, lamellar membranes associated percentages of GFP-LC3–positive cells were with DicsB in EM were also frequently seen quantified. *P G 0.0001. (G) atg5-knockout or Time after infection (min) normal MEF were infected with WT or DicsB for in the area at one end of the bacterial body, indicated periods. Intracellular multiplication of bacteria is determined. *P G 0.0001. Scale bars (A), which reminded us of the asymmetric (B), and (E), 2 mm. distribution of VirG (IcsA) on Shigella

728 4 FEBRUARY 2005 VOL 307 SCIENCE www.sciencemag.org R EPORTS required for actin-based intracellular motility VirGa1 in the presence of Atg5; Atg5-Myc VirG truncations, which suggests that they (15). We tested whether some autophagic was precipitated in a pull-down assay only share the amino acid residues 320 through component(s) might directly associate with when IcsB was absent (Fig. 3D). The 433 of VirG, which are involved in binding VirG and so possibly trigger the develop- extent of His-Atg5 binding to GST-VirGa1 (Fig. 3F). Indeed, in MDCK/pGFP-LC3 cells ment of autophagy around the bacterium. 1 decreased as we raised the amount of infected with DvirG/pD10-VirG2 (pVirG Cell lysates prepared from COS-7 cells purified IcsB added, which suggests that lacking VirG internal amino acid residues expressing GFP-Atg5, GFP-Beclin, GFP- Atg5 binding to VirG was inhibited by 319 to 507) but not DvirG/pVirG (full- LC3, or Myc-Vps34 were pulled down with IcsB in a dose-dependent manner (Fig. length VirG), the GFP-LC3 signals were GST-VirGa1 (the surface-exposed VirG 3E). To define the VirG region involved barely visible around bacteria (Fig. 3G). portion) (16), and the bound proteins were in binding, various truncated VirGa1 ver- Thus the interaction of IcsB with VirG by analyzed by immunoblotting. Note that only sions (VirGa1 through to VirGa8) (16) means of the putative VirG targeting se- Atg5 was reproducibly precipitated with were tested for the ability to bind to either quence for Atg5 might be a mechanism for GST-VirGa1 (fig. S6A). Furthermore, the His-Atg5 or IcsB by using the GST-VirG eluding the development of autophagy in binding capacity was also confirmed by per- pull down assay. Both His-Atg5 and IcsB the vicinity of the bacterial surface in mam- forming immunoprecipitation experiments werepulleddownbythesameseriesof malian cells. in 293T cells expressing Atg5-Myc and GFP-VirGa1 (fig. S6B), which raised the possibility that Atg5 might have some af- finity for VirG. To pursue this, BHK cells expressing GFP-Atg5 (BHK/pGFP-Atg5) were infected with DicsB for 4 hours and investigated for association of the bacterium with GFP-Atg5. The GFP-Atg5 signals were occasionally confined to one pole of the bacterium. It is noteworthy that when BHK/pGFP-Atg5 cells were infected with virG mutant (DvirG), the association of the Atg5 signal was barely detectable, but the signal was restored after infection with DvirG/pVirG (Fig. 3A). The Apg5 signal occasionally associated with VirG at one pole of the bacterium (Fig. 3B). To verify the concept, MDCK/pGFP-LC3 cells infected with WT, DicsB, DvirG, DvirG/ pVirG, or DvirG/DicsB (double mutant) were also examined for association of the LC3 signal with the bacteria. LC3-positive DvirG accounted for only 1.5%, whereas LC3-positive DicsB made up 37% (Fig. 3C). Although LC3-positive DvirG/pVirG was increased to 23%, exceeding that from WT (15%), LC3-positive DvirG/DicsB was still less than 2% (Fig. 3C). Consistently, similar results were obtained from the ex- periment using normal MEFs expressing GFP-LC3 (fig. S7, A and B). Then, we investigated the multiplicity of WT, DicsB, DvirG,andDvirG/DicsB within BHK cells. DvirG grew slightly better than WT within cells and found that the deficiency of intracellular growth of DicsB was recov- ered in DvirG/DicsB, which suggests that VirG is critical for Shigella to trigger autophagy (fig. S8). Atg5 may play a role as the Bseed[ of isolation membranes in the autophagic process (17). VirG present on Shigella could thus be the target for autophagy, in which IcsB would appar- ently act as the anti-autophagocytic agents. To assess this possibility, immobilized GST- VirGa1 with or without IcsB was incu- bated with a lysate of 293T cells expressing Fig. 2. Multilamellar structures enclosing DicsB in MDCK cells. (A) MDCK cells infected with DicsB Atg5-Myc, and the bound proteins were were observed by EM 4 hours later. Multilamellar structures surround the bacterium (arrows). (B) examined by immunoblotting. IcsB was ex- GFP-LC3 (arrows) is localized in the multilamellar structures (arrowheads) surrounding the amined in a pull-down assay with GST- bacterium, under immunogold EM. Scale bars, 0.5 mm.

www.sciencemag.org SCIENCE VOL 307 4 FEBRUARY 2005 729 R EPORTS

A B C 50 ) % ( ** 3

- 40 WT C L h t i

w 30 d e t a i c

o 20 s

s * a a i r e t 10 c a B B * ** ics ** 0 WT icsB virG virG/ virG/ icsB/ pVirG icsB pBR-IcsB Conc. of IcsB (µg/ml) D E

GST-VirG 1 t u 0 4 8 0 0 p 9 2

+Atg5-myc 9 0 . 6 3 . 5 . n 1 6 1 3 I 2 0 0 0 t B B u s s p / c B c I n I I

B -

+ His-Atg5 cs ics Atg5- Blot: -RGS-His

BR-I myc p Blot: -myc IcsB

IcsB Blot: -IcsB

Blot: -IcsB G

G F 1 2 3 4 5 6 7 8 ir

v G G G G G G G G r r r r r r r r i i i i i i i i G G

ir ir V V V V V V V V t ------v V u T T T T T T T T T /p p S S S S S S S S S n I G G G G G G G G G

His- Atg5 Blot: -RGS-His

IcsB G2 G ir / ir

v

G Blot: -IcsB G

ir ir

v V 53 759 1102 D10-V

p Binding N -domain -core C /p Atg5 IcsB 53 779 GST- 1 + + 103 320 GST- 2 + + 506 GST- 3 + + 319 GST- 4 - - 320 507 GST- 5 + + 101 GST- 6 - - 103 433 GST- 7 + + 320 433 GST- 8 + +

Fig. 3. VirG affinity for Apg5 is inhibited by IcsB. (A) BHK/pGFP-Atg5 cells GST-VirGa1 with the use of cleared lysates of 293T cells expressing infected with indicated Shigella strains for 4 hours were stained with antibody Atg5-Myc. (E) Dose-response analysis of IcsB inhibition for Atg5 binding against Shigella (red). (B) BHK/pGFP-Atg5 cells infected with DvirG/pVirG for to GST-VirGa1. (F) Determination of VirG domain involved in binding 4 hours were stained with antibody against VirG (red). (C)MDCK/pGFP-LC3 with Atg5 and IcsB (top) and schematic representation of the virGa1 cells infected with indicated Shigella strains for 4 hours were stained with derivatives (bottom). (G) MDCK/pGFP-LC3 cells infected with DvirG/pVirG antibody against Shigella. The percentages of GFP-LC3–positive bacteria were or DvirG/pD10-VirG2 for 4 hours were stained with antibody against quantified. *P G 0.0001. (D) Analysis of IcsB inhibition for Atg5 binding to Shigella (red). Scale bars (A), (B), and (G), 2 mm.

To demonstrate whether VirG itself was a association with LC3 was investigated. In Shigella VirG can thus be targeted for target for autophagy, BHK cells expressing cells infected with E. coli/pSU-GFP/pUC- autophagy perhaps through its affinity for Myc-LC3 (BHK/pMyc-LC3) were infected VirG but not E. coli/pSU-GFP, bacterial Atg5, and IcsB can interfere with this with an E. coli K-12 strain expressing VirG signals were localized with Myc-LC3 sig- autophagic process. The role of IcsB in plus GFP (E. coli/pSU-GFP/pUC-VirG) or GFP nals, and the VirG-associated signals merged bacterial infection differs from other known (E. coli/pSU-GFP) together with DvirG/DicsB with the GFP-LC3 signals (Fig. 4). It is thus TTSS-secreted bacterial effectors, in that its (invasion-positive but autophagy-negative highly likely that VirG presented on the role is to camouflage its own bacterial target S. flexneri used as the carrier, which allowed Shigella surface in the host cytoplasm trig- molecule (VirG) from the autophagic host E. coli to move into the host cytoplasm). The gers autophagy. defense system.

730 4 FEBRUARY 2005 VOL 307 SCIENCE www.sciencemag.org R EPORTS

Priority Areas from the Ministry of Education, Culture, Figs. S1 to S8 Sports, and Technology (MEXT). Molecular interaction Table S1 data have been deposited in the Biomolecular Inter- References action Network Database with accession code 185200. 5 October 2004; accepted 19 November 2004 /

P Supporting Online Material Published online 2 December 2004;

F www.sciencemag.org/cgi/content/full/1106036/DC1 10.1126/science.1106036 G G r

- Materials and Methods Include this information when citing this paper. i U V - S C p / i

l U

o p

c . Nod2-Dependent Regulation of E Innate and Adaptive Immunity in the Intestinal Tract Koichi S. Kobayashi,1* Mathias Chamaillard,3,4 Yasunori Ogura,1 P

/ 1 3 3,4 i F . l Octavian Henegariu, Naohiro Inohara, Gabriel Nun˜ez,

o G

- 1,2 c .- . Richard A. Flavell U

E S p The gene encoding the Nod2 protein is frequently mutated in Crohn’s disease (CD) patients, although the physiological function of Nod2 in the intestine remains elusive. Here we show that protective immunity mediated by Nod2 recognition of bacterial muramyl dipeptide is abolished in Nod2-deficient mice. These animals are susceptible to bacterial infection via the oral route but not through intravenous or peritoneal delivery. Nod2 is required for the expression of a subgroup of intestinal anti-microbial peptides, known as cryptdins. The G / r

i i l Nod2 protein is thus a critical regulator of bacterial immunity within the

o V - c intestine, providing a possible mechanism for Nod2 mutations in CD. . C

E U

p Homozygous mutations in Nod2, a member symptoms of intestinal inflammation when ob- of the nucleotide-binding oligomerization served for up to 6 months, which was Fig. 4. VirG triggers autophagy. BHK/pMyc-LC3 domain–leucine-rich repeat (NOD-LRR) confirmed by histological examination (fig. coinfected with E. coli/pSU-GFP/pUC-VirG and family of proteins, are highly correlated with S3), and there was no significantly enhanced DvirG/DicsB at a ratio of 10:1 for 2 hours were the incidence of Crohn_sdisease(CD),which susceptibility to colitis in the dextran sulfate– stained with antibody against Myc (red) (top). As a suggests that Nod2 plays an important role in induced model (fig. S4). control, E. coli/pSU-GFP was used. (Bottom)BHK/ intestinal immunity (1, 2). These CD-associated Detection of MDP by Nod2 was con- pGFP-LC3 cells were coinfected with E. coli/pUC- VirG and DvirG/DicsB as described above, and genetic variants are deficient in their ability to firmed by stimulation of wild-type and j/j stained with antibody against VirG (red) (bottom). sense muramyl dipeptide (MDP), a conserved Nod2 bone marrow–derived macrophages Scale bar, 2 mm. Note that as E. coli/pUC-VirG structure in bacterial peptidoglycan that is nor- (BMDMs) with the following Toll-like re- expressed a high level of surface VirG, the VirG mally recognized by Nod2 (3–5). Consistent ceptor (TLR) agonists: lipopolysaccharide signal was visible over the entire bacterial surface. with these functional studies, homozygosity and (LPS, TLR4 ligand); Pam3CS(K)4 (synthetic compound heterozygosity increase the relative lipopeptide, TLR2 ligand); and double- References and Notes risk of developing CD by as much as 40-fold as stranded RNA Epolyinosinic:polycytidylic 1. K. A. Rich, C. Burkett, P. Webster, Cell. Microbiol. 5, compared to simple heterozygosity (È2- to 4- acid (poly I:C), TLR3 ligand^ in the presence 455 (2003). 2. B. R. Dorn, W. A. Dunn Jr., A. Progulske-Fox, Cell. fold) (1, 2, 6). Nevertheless, the physiological or absence of MDP (8–12). The synergistic Microbiol. 4, 1 (2002). role of Nod2 in intestinal immunity remains effect of MDP and TLR ligands for the pro- 3. S. Me´resse et al., Nature Cell Biol. 1, E183 (1999). unclear. duction of interleuken-6 (IL-6) and IL-12 4. L. D. Hernandez, M. Pypaert, R. A. Flavell, J. E. Ga´lan, J. Cell Biol. 163, 1123 (2003). To assess the function of Nod2, we gener- p40 in wild-type macrophages was depen- j j 5. K. Kirkegaard, M. P. Taylor, W. T. Jackson, Nature Rev. ated Nod2 / mice using a targeting construct dent on the dose of MDP added (Fig. 1, A Microbiol. 2, 301 (2004). to replace the NOD, which is essential for and B, and fig. S5F) and was absent in 6. G. Marin˜o, C. Lo´pez-Otı´n, Cell. Mol. Life Sci. 61, 1439 j/j (2004). activation of the protein (fig. S1, A and B) (7). Nod2 macrophages, revealing that Nod2 j j 7. A. M. Cuervo, Trends Cell Biol. 14, 70 (2004). Nod2 / mice were outwardly healthy and is required for the detection of MDP in 8. M. L. Bernardini et al., Proc. Natl. Acad. Sci. U.S.A. displayed normal lymphoid and myeloid cel- macrophages. The stimulation of wild-type 86, 3867 (1989). 9. P. Cossart, P. J. Sansonetti, Science 304, 242 (2004). lular composition in the thymus and spleen macrophages with MDP, which results in the 10.L.G.Tilney,D.A.Portnoy,J. Cell Biol. 109, 1597 (1989). (fig. S2). The animals also displayed no overt activation of nuclear factor kB (NF-kB), p38, 11. E. Gouin et al., Nature 427, 457 (2004). 1 2 and extracellular signal–regulated kinase 12. M. Ogawa et al., Mol. Microbiol. 48, 913 (2003). Section of Immunobiology, The Howard Hughes (ERK), was strongly reduced in Nod2–/– 13. Y. Kabeya et al., EMBO J. 19, 5720 (2000). Medical Institute, Yale University School of Medicine, 14. T. Yoshimori, Biochem. Biophys. Res. Commun. 313, New Haven, CT 06510, USA. 3Department of Pathol- macrophages, again indicating that Nod2 is 453 (2004). ogy, 4Comprehensive Cancer Center, The University of required for MDP signaling (Fig. 1C). In light 15. J. R. Robbins et al., Mol. Microbiol. 41, 861 (2001). Michigan Medical School, Ann Arbor, MI 48109, USA. of the expression of Nod2 in CD40þ/CD86þ 16. T. Suzuki, S. Saga, C. Sasakawa, J. Biol. Chem. 271, 21878 (1996). *Present address: Dana-Farber Cancer Institute and dendritic cells (DCs) (13), we tested the 17. N. Mizushima et al., J. Cell Biol. 152, 657 (2001). Harvard Medical School, 44 Binney Street, Boston, potential involvement of the protein in the 18. We thank members of the Sasakawa laboratory for MA 02115, USA. detection of MDP by DCs. Although im- technical advice and experimental support. We thank .These authors contributed equally to this work. R. Akakura for critical reading of the manuscript. This work -To whom correspondence should be addressed. mature bone marrow–derived dendritic cells was supported by a grant-in-aid for Scientific Research on E-mail: [email protected] (BMDCs) from wild-type mice were capable

www.sciencemag.org SCIENCE VOL 307 4 FEBRUARY 2005 731 R EPORTS of secreting the proinflammatory cytokines and IgG1 at a similar level to wild-type mice 3A). Similarly, serum IL-6 production, which IL-6 and –a (TNF-a), when they were immunized with resiquimod normally accompanies bacterial infection, these cytokines were not detectable from (R-848, a synthetic ligand for mouse TLR7) was not elevated. (Fig. 3B). Because L. Nod2–/– immature BMDCs (Fig. 1D). MDP (Fig. 2, E and F) (17). These results indicate monocytogenes naturally infects humans by alone did not induce cytokine production that Nod2 is able to activate adaptive immu- the intestinal route, mice were also chal- from BMDMs. nity and to mediate adjuvant activity in the lenged with L. monocytogenes by intra- Previous studies have shown that pre- production of antibody to T cell–dependent peritoneal injection, and again no significant treatment with MDP sensitizes mice to . difference in survival between wild-type and endotoxic shock induced by LPS injection To investigate whether Nod2 plays a spe- Nod2j/j mice was seen (Fig. 3C). Consist- (14). Although pretreatment with 250 mgof cific role in the innate immune response ent with these data, in vitro infection of MDP and subsequent highly purified LPS against bacterial infection, wild-type and macrophages from Nod2j/j mice with L. injection were fatal in wild-type mice by day Nod2j/j mice were challenged with the monocytogenes induced equivalent level of 6, Nod2j/j mice were resistant to LPS Gram-positive intracellular bacterium Liste- TNF-a and resulted in similar intracellular challenge (Fig. 1E). LPS injection without ria monocytogenes by intravenous injection. bacterial growth and killing (fig. S6, A to C). MDP priming resulted in the similar survival No significant difference was observed in the In contrast, Nod2j/j mice challenged with L. of both wild-type and Nod2j/j animals. number of bacteria recovered from both the monocytogenes via intragastric dosing were These indicate an essential role for Nod2 in liver and the spleen 24 or 48 hours later (Fig. susceptible to infection and showed signifi- detecting MDP in vivo. Nod2 has recently been proposed to serve AB40 as a negative regulator of TLR2 signaling in WT 60 35 WT Nod2-/- generating the Th1 phenotype (15). In testing 50 j j 30 Nod2-/-

/ ml) ml) /

the response of Nod2 BMDMs for the / g g

25 n production of inflammatory cytokines, in- n 40

20 40 (

cluding IL-6, IL-12 p40, TNF-a, and IL-1b, p IL-6 ( 30 after stimulation of various TLR ligands, 15

IL-12 20 including TLR2 ligands (fig. S5, A to E), we 10 failed to observe any significant diminution 5 10 in the response of Nod2j/j macrophages to 0 0 either LPS and TLR2 ligands (fig. S5, A to MDP (µg/ml) 011001 10 MDP (µg/ml) 011001 10 C). We also observed a synergistic effect of Poly(IC) LPS Poly(IC) LPS lipopeptide Pam3CS(K)4 (TLR2 ligand) and MDP in wild-type macrophages but not in C WT Nod2-/- D j/j 020601010 02060min 3.0 7.0 Nod2 macrophages (fig. S5F). These MDP MDP κ α untreated untreated results suggest that any potential negative pI B 2.5 6.0 ml)

role of Nod2 in the TLR2 response is not a κ α / 5.0 I B ml) / 2.0 g g universal phenomenon. Determination of n n ( 4.0 whether there is a contribution to TLR2 1.5 α pp38 3.0

response from specific experimental details, IL-6 ( p38 1.0 TNF- genetic background, or cell types examined 2.0 0.5 will require further study. pERK1/2 1.0 In light of previous studies showing the 0.0 0.0 role of TLRs in the adaptive immune system ERK1/2 WT Nod2-/- WT Nod2-/- (16), we immunized wild-type and Nod2j/j mice with MDP (100 mg per mouse) and with E + MDP - MDP human serum albumin (HSA) (50 mg per 100 100 mouse) by intraperitoneal injection. Three weeks later, we boosted with MDP (20 mg 80 80 per mouse) and HSA (10 mg per mouse). 60 60 val (%) Serum samples were obtained 2 weeks after i v r the first immunization and 1 week after the 40 40 Su boost, and -specific serum immuno- WT globulin (Ig) was assessed by enzyme-linked 20 20 Nod2-/- immunosorbent assay (ELISA). Although 0 0 HSA-specific antibodies were detected 2 days 0691423 5 78 10 0691423 5 78 10 weeks after immunization in wild-type mice, Nod2j/j animals displayed a severe defi- Fig. 1. Nod2 cooperates with TLR signaling and is required for MDP recognition in vivo. (A) BMDMs from wild-type (WT) and Nod2j/j mice were stimulated with LPS (10 ng/ml) (from Alexis) or ciency in the production of antigen-specific poly(I:C) (100 mg/ml) in the presence of MDP (0, 1, or 10 mg/ml) for 24 hours. Production of IL-6 Ig, specifically in IgG1, which is the pre- was assessed by ELISA. (B) Production of IL-12 p40 was assessed as in (A). (C) BMDMs from wild- dominant Nod2-dependent isotype (Fig. 2, type and Nod2j/j mice were stimulated with MDP (10 mg/ml) for indicated periods. Total cell A and B). The defect in antigen-specific Ig lysates were prepared and blotted with antibodies against phosphorylated (p) and unphosphoryl- ated forms of IkBa, p38, and ERK1/2. (D) Immature BMDCs were stimulated with MDP (25 mg/ml) production was observed even after boosting 0 mice with MDP and HSA. Nod2j/j mice for 20 hours, and production of IL-6 and TNF-a was assessed by ELISA. (E) Wild-type (n 8) and Nod2j/j (n 0 6) mice (8 to 12 weeks old) were primed with MDP (250 mg) and challenged with failed to produce HSA-specific Ig and IgG1 m j j LPS (250 g, Ultrapure LPS from InvivoGen). The survival of each mouse genotype was plotted. As (Fig. 2, C and D). In contrast, Nod2 / mice a control, wild-type (n 0 11) and Nod2j/j (n 0 8) mice were challenged with LPS without MDP were capable of producing HSA-specific Ig priming.

732 4 FEBRUARY 2005 VOL 307 SCIENCE www.sciencemag.org R EPORTS Fig. 2. Impaired antigen- AB35 cantly greater numbers of bacteria recovered specific immunoglobulin pro- 400 from both the liver and the spleen than did j/j 30 duction in Nod2 mice 350 ml) / wild-type mice (Fig. 3D). These results ml) g / immunizedwithMDP.(A) n g 300 25 Wild-type (n 0 5) and n suggest that Nod2 plays a pivotal and spe- j/j 0 250 cific role in protecting against bacterial in- Nod2 (n 4) mice (8 Ig ( 20 IgG1 ( ic f

ic WT fection in the intestine. The number of L. to 12 weeks old) were 200 f immunized with HSA (100 WT 15 Nod2-/- monocytogenes in intestinal Peyer_s patches speci 150 Nod2-/- mg) and MDP (50 mg) and speci 10 was not significantly different between wild- bled 2 weeks after immu- 100 –/– HSA- type and Nod2 mice, which suggests that nization. Antigen-specific 50 HSA- 5 _ Ig was measured by ELISA. Nod2 might be involved in a Peyer s patch– The P value was deter- 0 0 independent route of bacterial invasion (fig. P<0.005 mined by Student’s t-test. P<0.005 S7). Expression of Nod2 in the terminal ileum (B)Antigen-specificIgG1 CD20 4.5 was examined by using purified villi and was assessed by ELISA as 18 4.0 crypts (Fig. 3E) (18). Reverse transcription

in (A). (C) The immunized ml) / ml) polymerase chain reaction (RT-PCR) revealed / 16 g 3.5 g mice were boosted 3 weeks µ µ 14 that Nod2 is highly expressed in crypts but not later with HSA (20 mg) and 3.0

MDP (10 mg) and bled 1 Ig ( 12 in villi, which is consistent with a recent re- ic 2.5 IgG1 ( f WT WT week later. Antigen-specific 10 ic port showing expression in human intestinal Nod2-/- f 2.0 Nod2-/- Ig was measured by ELISA. 8 Paneth cells (Fig. 3F) (19, 20). speci 1.5 (D)Antigen-specificIgG1 6 speci To investigate potential genes that might was assessed by ELISA as HSA- 4 1.0 be induced by Nod2 during intestinal infec- in (C). (E)Wild-type(n 0 5) HSA- 2 0.5 and Nod2j/j (n 0 4) mice tion, we isolated RNA samples from both the 0 0 wild-type and Nod2j/j terminal ileum be- (8 to 12 weeks old) were P<0.005 P<0.005 immunized with HSA (100 fore and after Listeria infection by gastric mg) and R-848 (100 nmol) EF2.5 gavage and screened them by microarray and bled 2 weeks after im- analysis (table S2). The up-regulation of 2.0 1.2 munization. Antigen-specific ml) / ml)

/ candidate genes, which might explain the g g

Ig was measured as in (A). n 1.0 j j n susceptibility of Nod2 / mice to bacterial (F)Antigen-specificIgG1 1.5 Ig ( 0.8 infection, was confirmed by quantitative ic

was assessed by ELISA as f

IgG1 ( WT in (E). WT 0.6 real-time PCR (Fig. 3, G to I). The most ic 1.0 f Nod2-/- speci Nod2-/- significant difference was in the expression 0.4 0.5 speci of a subgroup of cryptdins (called a- HSA- 0.2 defensins in humans). Thus, expression of HSA- 0 0 defensin-related cryptdin 4 (Defcr4) and

Fig. 3. Nod2 is required for the expression of cryptdins and for protection animal and bars indicate the mean value. (E) Preparation of mouse small against intracellular bacterial infection in the intestine. (A) Wild-type and intestinal crypts and villi. Individual mouse crypts were stained for 2 min Nod2j/j mice (8 to 12 weeks old) were infected with 3 103 L. with 0.25% amido black (marked by an arrow). (F) Villi and crypts were monocytogenes (strain 10403S) intravenously, and the number of bacteria isolated from the terminal ileum of wild-type and Nod2j/j mice, and the in the liver and the spleen was counted 24 and 48 hours after infection. expression of Nod2 was examined by RT-PCR. Glyceraldehyde phosphate (B) Wild-type (n 0 5) and Nod2j/j (n 0 5) mice were infected with 104 L. dehydrogenase (GAPDH) and Defcr3 were used for loading control and monocytogenes intravenously and serum IL-6 was measured by ELISA. (C) quality of the crypt preparation, respectively. (G to I) Wild-type and Wild-type (n 0 14) and Nod2j/j (n 0 13) mice were infected with 106 L. Nod2j/j mice (8 to 12 weeks old) were infected with 109 L. monocytogenes intraperitoneally and the survival of the mice was plotted. monocytogenes intragastrically by gastric gavage. mRNA was extracted (D) Wild-type (n 0 10) and Nod2j/j (n 0 10) mice were infected with 109 from the terminal ileum from Nod2j/j mice before (n 0 5) and 24 hours L. monocytogenes intragastrically by gastric gavage, and the number of after (n 0 4) infection and from littermate wild-type mice before (n 0 9) bacteria in the liver and the spleen was counted 72 hours after infection. and 24 hours after (n 0 4) infection. The expression of Defcr4, Defcr-rs10, The P value was determined by Mann-Whitney test. Circles indicate each and Defcr5 was examined by quantitative real-time PCR.

www.sciencemag.org SCIENCE VOL 307 4 FEBRUARY 2005 733 R EPORTS Defcr-related sequence 10 (Defcr-rs10) was mutations predispose primarily to ileal lesions 18. T. Ayabe et al., Nature Immunol. 1, 113 (2000). very low in Nod2j/j mice and was further (30–33), corresponding to the location of 19. S. Lala et al., Gastroenterology 125, 47 (2003). 20. Y. Ogura et al., Gut 52, 1591 (2003). j/j reduced after infection in Nod2 animals Paneth cells. Recent reports suggest that the 21. P. B. Eisenhauer, S. S. Harwig, R. I. Lehrer, Infect. relative to wild-type mice (Fig. 3, G and H). expression of a-defensins is diminished in Immun. 60, 3556 (1992). Cryptdins are antimicrobial peptides that are human CD patients, particularly those who 22. S. A. Naser, G. Ghobrial, C. Romero, J. F. Valentine, Lancet 364, 1039 (2004). preferentially produced in intestinal Paneth have Nod2 gene mutations (34, 35). However, 23. W. S. Selby, Lancet 364, 1013 (2004). cells, and their antimicrobial activity is im- it remains to be established whether a defect 24.A.J.Ouelletteet al., Infect. Immun. 62, 5040 portant in suppressing infection with patho- in Paneth cell function is the only possible (1994). 25.A.J.Ouelletteet al., Infect. Immun. 67, 6643 genic bacteria, including L. monocytogenes mechanism by which Nod2 mutations might (1999). (21) and Mycobacterium paratuberculosis, associate with the development of CD in 26. D. Darmoul, A. J. Ouellette, Am. J. Physiol. 271, G68 an organism implicated in CD (22, 23). Of humans. Nevertheless, it seems reasonable (1996). 27. K. Tani et al., Int. Immunol. 12, 691 (2000). the cryptdins, Defcr4 has the most potent to suggest that mutations in Nod2 might 28. D. Yang, A. Biragyn, L. W. Kwak, J. J. Oppenheim, bactericidal activity (24), with its expression promote CD through defective regulation of Trends Immunol. 23, 291 (2002). being highest in the lower ileum, in con- responses to commensal and/or pathogenic 29. C. L. Wilson et al., Science 286, 113 (1999). 30. T. Ahmad et al., Gastroenterology 122, 854 (2002). trast to other cryptdins (25, 26). By com- bacteria, rather than acting as an initiating 31. A. P. Cuthbert et al., Gastroenterology 122, 867 parison, Defcr5 was expressed normally in factor for disease. Further studies may re- (2002). Nod2j/j mice both before and after infec- solve this issue and may lead to the develop- 32. S. Lesage et al., Am. J. Hum. Genet. 70, 845 (2002). tion (Fig. 3I) (25). ment of more rational therapeutic approaches 33. J. Hampe et al., Lancet 359, 1661 (2002). 34. M. Schmid, K. Fellermann, J. Wehkamp, K. Herrlinger, Our results indicate that Nod2 is essen- for treating CD. E. F. Stange, Z. Gastroenterol. 42, 333 (2004). tial in the detection of bacterial MDP and is 35. J. Wehkamp et al., Gut 53, 1658 (2004). capable of activating the adaptive immune References and Notes 36. The authors thank C. L. Stewart for providing 1. Y. Ogura et al., Nature 411, 603 (2001). reagents; L. Evangelisti, M. Chen, and A. Ferrandino system by acting as an adjuvant receptor for 2. J. P. Hugot et al., Nature 411, 599 (2001). for technical assistance; and F. Manzo for manu- antibody production, either directly or by 3. N. Inohara et al., J. Biol. Chem. 278, 5509 (2003). script preparation. Supported by grants from NIH enhancing the production of a-defensins 4. S. E. Girardin et al., J. Biol. Chem. 278, 8869 (2003). (K.S.K., N.I., G.N., and R.A.F.), the Crohn’s and Colitis 5. J. Li et al., Hum. Mol. Genet. 13, 1715 (2004). Foundation of America (K.S.K., M.C., Y.O., and R.A.F.), (27, 28) or other immunostimulatory mole- 6. J. Hampe et al., Lancet 357, 1925 (2001). the Ellison Foundation (R.A.F.), and the Eli and cules. Therefore, Nod2 is critical in protecting 7. Y. Ogura et al., J. Biol. Chem. 276, 4812 (2001). Edythe L. Broad Foundation (K.S.K. and G.N.). R.A.F. the host from intestinal bacterial infection. 8. A. Poltorak et al., Science 282, 2085 (1998). is an investigator of the Howard Hughes Medical More specifically, we reveal an important role 9. S. T. Qureshi et al., J. Exp. Med. 189, 615 (1999). Institute. 10. K. Hoshino et al., J. Immunol. 162, 3749 (1999). for Nod2 in the regulation of a subgroup of 11. A. O. Aliprantis et al., Science 285, 736 (1999). Supporting Online Material cryptdins, offering a plausible mechanism to 12. L. Alexopoulou, A. C. Holt, R. Medzhitov, R. A. Flavell, www.sciencemag.org/cgi/content/full/307/5710/731/ explain the association between Nod2 and Nature 413, 732 (2001). DC1 13. O. Gutierrez et al., J. Biol. Chem. 277, 41701 (2002). Materials and Methods susceptibility to CD. Murine cryptdins repre- 14. H. Takada, S. Yokoyama, S. Yang, J. Endotoxin Res. 8, Figs. S1 to S7 sent a more diverse family than those of hu- 337 (2002). Tables S1 and S2 man a-defensins and are already known to be 15. T. Watanabe, A. Kitani, P. J. Murray, W. Strober, References Nature Immunol. 5, 800 (2004). critical in the innate immune responses to 16. M. Schnare et al., Nature Immunol. 2, 947 (2001). 7 September 2004; accepted 9 December 2004 bacterial infection (29). CD-associated Nod2 17. H. Hemmi et al., Nature Immunol. 3, 196 (2002). 10.1126/science.1104911

main (NBD), and 10 C-terminal leucine-rich Nod2 Mutation in Crohn’s Disease repeats (LRRs), and it is expressed mainly k by macrophages and dendritic cells (5). Potentiates NF- B Activity NOD2 mediates intracellular recognition of MDP, a building block for bacterial cell and IL-1b Processing walls (6, 7), and can activate NF-kB(5). Macrophages within the intestinal lamina Shin Maeda,1 Li-Chung Hsu,1* Hongjun Liu,1* propria of CD patients overproduce NF-kB Laurie A. Bankston,1,3 Mitsutoshi Iimura,2 Martin F. Kagnoff,2 targets, including the proinflammatory cyto- Lars Eckmann,2 Michael Karin1. kines tumor necrosis factor–a (TNFa) and the interleukins IL-1b and IL-6 (2, 8). Many Variants of NOD2, an intracellular sensor of bacteria-derived muramyl di- of the anti-inflammatory drugs used to treat peptide (MDP), increase susceptibility to Crohn’s disease (CD). These variants CD inhibit NF-kB activation, which sug- are thought to be defective in activation of nuclear factor kB (NF-kB) and gests it is a key pathogenic factor (8, 9). antibacterial defenses, but CD clinical specimens display elevated NF-kB However, paradoxically, transient transfec- activity. To illuminate the pathophysiological function of NOD2, we intro- tion experiments indicate that CD-associated duced such a variant to the mouse Nod2 locus. Mutant mice exhibited k elevated NF- B activation in response to MDP and more efficient processing 1Laboratory of Gene Regulation and Signal Transduc- and secretion of the cytokine interleukin-1b (IL-1b). These effects are linked tion, Department of Pharmacology and 2Laboratory to increased susceptibility to bacterial-induced intestinal inflammation and of Mucosal Immunology, Departments of Medicine identify NOD2 as a positive regulator of NF-kB activation and IL-1b secretion. and Pediatrics, School of Medicine, University of California, San Diego, 9500 Gilman Drive, La Jolla, 3 Crohn_s disease (CD) is a chronic inflamma- Mutations that increase susceptibility to CD CA 92093–0723, USA. Program on Cell Adhesion, The Burnham Institute, 10901 North Torrey Pines tory bowel disease (IBD) thought to be up to 40 times were mapped to the NOD2/ Road, La Jolla, CA 92037, USA. caused by genetic and environmental factors CARD15 locus (3, 4). The NOD2 protein *These authors contributed equally to this work. that affect host-microbe interactions and pro- contains two N-terminal caspase recruitment .To whom correspondence should be addressed. duction of inflammatory mediators (1, 2). domains (CARDs), a nucleotide-binding do- E-mail: [email protected]

734 4 FEBRUARY 2005 VOL 307 SCIENCE www.sciencemag.org R EPORTS

Fig. 1. Generation of Nod22939iC mice. (A) Schematic structure of NOD2, sequence of WT, and mutant alleles around the 2939insC mutation, targeting vector, and the targeted locus. Solid boxes repre- sent exons, and lines introns. The Neor cassette was inserted opposite the Nod2 transcription unit. (B) Southern blot analy- sis of Nco I–digested

genomic DNA from F2 mice of the in- dicated genotypes, where m is mutant allele; þ, WT allele. (C) Nod2 mRNA in BMDMs. RNA was converted to cDNA and amplified by using primers for three dif- ferent regions of Nod2 cDNA. (D) Expres- sion of WT and truncated (m/m) NOD2 proteins. BMDM lysates were immuno- blotted with antibodies against NOD2 and actin in order to control loading.

NOD2 variants no longer activate NF-kB cultures. The activity of IKK, which is the microbial product that stimulated IL-1b se- in response to MDP (6, 7), which suggests inhibitor of kB(IkB) kinase, the degradation cretion by Nod22939iC macrophages was MDP that defective NF-kB activation in macro- of IkBa, and NF-kB DNA binding activity (Fig. 2E). phages facilitates infection of the lamina were higher in MDP-stimulated Nod22939iC Macrophages involved in CD most likely propria by enteric bacteria. However, mac- macrophages than in wild-type (WT) cells reside in the lamina propria (2). To expose rophages can activate NF-kB in response (Fig. 2A). Only marginal differences in these cells to enteric bacteria, mice were to bacteria independently of NOD2 (10), mitogen-activated protein kinases (MAPKs) treated with dextran sodium sulfate (DSS), and Nod2 gene ablation did not cause spon- were observed (fig. S2). No genotype-specific an agent that kills mucosal epithelial cells taneous intestinal infections or colonic in- differences in NF-kB activation were ob- and disrupts their barrier function, causing flammation (11). served after macrophage treatment with bacterial invasion (13). WT and homozygous To find an explanation for these quanda- other microbial components that activate Nod22939iC mice (8 to 12 weeks old) were ries and to illuminate the mechanism by Toll-like receptors (TLRs) (10), including given 3% DSS in drinking water for 6 days which CD-associated NOD2 variants act, we the TLR2 agonists Pam3Cys (tripalmitoyl-S- and monitored for weight loss, a charac- generated mice whose Nod2 locus harbors glyceryl-Cys-Ser-4(Lys)) and peptidoglycan teristic of severe intestinal inflammation. the homolog of the most common CD sus- (PGN), the TLR4 agonist lipopolysaccha- After 8 days, body weight loss was greater ceptibility allele, 3020insC, which encodes a ride (LPS), and the TLR9 agonist nonmeth- in Nod22939iC mice relative to WT mice truncated protein lacking the last 33 amino ylated CpG-containing DNA (Fig. 2B) (12). (Fig. 3A). Nod22939iC mice also exhibited in- acids (3, 4). This was done through insertion Expression of several NF-kB target genes creased mortality relative to WT mice (37.5% of cytosine at position 2939 (corresponding was increased in MDP-treated Nod22939iC versus 0%) (fig. S4). Surviving mice of both to 3020 in human NOD2) of the Nod2 open macrophages relative to WT counterparts genotypes regained body weight after day reading frame (Fig. 1, A and B). Homo- (Fig. 2C). Only minor differences in expres- 11 and returned to normal 30 days after DSS zygous Nod22939iC mice were obtained at the sion of these genes were observed when administration (12). Histological analyses expected mendelian ratio and did not show macrophages were stimulated with LPS or revealed that the severity and extent of inflam- abnormalities of the gastrointestinal tract PGN. Although MDP-induced gene expres- matory lesions in the colons of Nod22939iC (fig. S1) or other organs; they were healthy sion of several cytokine genes was increased mice were significantly (P G 0.05) greater (12). The mutation had no effect on Nod2 in Nod22939iC macrophages, only IL-1b se- than in WT controls, with larger areas of ul- mRNA or protein amounts in bone marrow– cretion was significantly elevated in these ceration and increased infiltration of F4/80- derived macrophages (BMDMs) (Fig. 1, C cells relative to WT counterparts (Fig. 2, D positive macrophages (Fig. 3B, fig. S5). and D). and E, fig. S3). Secretion of IL-1a was mod- After DSS exposure, Nod22939iC homozy- We examined the effect of the Nod22939ic estly elevated, and neither IL-6 nor TNFa gotes expressed greater amounts of mRNAs mutation on NF-kB activation in BMDM were secreted in response to MDP. The only encoding proinflammatory cytokines and che-

www.sciencemag.org SCIENCE VOL 307 4 FEBRUARY 2005 735 R EPORTS mokines in their colons relative to WT mice to DSS (fig. S11). However, its deletion in product induces elevated IKK and caspase-1 (Fig. 3C). The amounts of IL-1b,IL-6,and enterocytes increased the inflammatory re- activation in response to MDP. Although cyclooxygenase-2 (Cox-2) protein were signif- sponse to DSS (19). NOD2 was suggested to be a negative reg- icantly higher in colons of DSS-treated Collectively, our results suggest that ulator of TLR2 (20), we found no effect of Nod22939iC mice relative to WT counterparts Nod22939iC is a gain-of-function allele, whose the Nod22939iC mutation on signaling by TLR2, (Fig. 3D). IL-6 and Cox-2 were predominantly expressed in F4/80-positive macrophages within inflammatory lesions (Fig. 3E, fig. S6) (12). IKK and NF-kB activities and RelA(p65) nuclear staining were also higher in colons of Nod22939iC mice than in the WT (Fig. 3F, fig. S7). MAPK activation, how- ever, was only marginally affected by the genotype (fig. S8). The intestinal inflammatory response to DSS is dramatically reduced by oral anti- biotics, which supports involvement of en- teric bacteria (14). When given a high dose of DSS (6%) without oral antibiotics, WT and Nod22939iC mice died within 9 days after DSS administration (12), but mice that received oral antibiotics survived and devel- oped mild inflammation and weight loss, without any genotype-linked differences (fig. S9). Thus, enteric bacteria elicit the inflam- matory response to DSS, and without bacte- rial exposure, Nod22939iC mice have the same reaction as WT counterparts. Exposure of macrophages to bacteria acti- vates inflammatory and apoptotic caspases (15). More apoptotic cells, most of which were positive for the F4/80 macrophage marker, were found in the lamina propria of DSS-treated Nod22939iC mice than in WT counterparts (Fig. 4, A and B). Increased macrophage apoptosis is associated with ac- tivation of caspase-1 (16), an enzyme required for secretion of mature IL-1b (17, 18). Congruently, only background levels of se- creted IL-1b were present in colons of un- treated mice, but IL-1b concentrations were elevated after DSS treatment, particularly in Nod22939iC mice (Fig. 3D). Macrophage acti- vation with LPS induces pro–IL-1b, but its processing and release require activation of caspase-1 by a different signal (16). LPS did not induce secretion of mature IL-1b in either Nod22939iC or WT macrophages, although it stimulated TNFa release (Fig. 2, D and E). In

contrast, MDP stimulated release of mature 2939iC 2939iC Fig. 2. Nod2 macrophages exhibit elevated NF-kB activation and IL-1b secretion in response IL-1b but not TNFa by Nod2 macro- to MDP. (A) BMDMs from WT and Nod22939iC (m/m) mice were incubated with MDP (1 mg/ml). phages. To determine whether IL-1b secretion Where indicated, cytosolic and nuclear extracts were prepared and used to analyze IKK activation may be involved in the increased inflamma- (KA), IkBa degradation, and NF-kB DNA binding activity, respectively. Nuclear extract quality was tory response to DSS in Nod22939iC mice, monitored by measuring nuclear factor-Y (NF-Y) DNA binding. (B) BMDMs were stimulated with mice were injected once daily with IL-1 Pam3Cys (1 mg/ml), LPS (100 ng/ml), or CpG DNA (1 mM) to activate TLR2, 4, and 9, respectively. receptor antagonist (IL-1-RA) from the start Where indicated, nuclear extracts were prepared and NF-kB DNA binding activity was analyzed. (C) Expression of NF-kB target genes was examined in Nod22939iC and WT macrophages stimulated of DSS exposure. Average body weight loss with MDP, LPS, or peptidoglycan (PGN from Staphylococcus aureus,10mg/ml). After 4 hours, cells and histological score were improved in IL-1- were collected, total cytoplasmic RNA was prepared, and gene expression was analyzed by real- RA–treated mice, and differences in weight time polymerase chain reaction (PCR). Data are presented as the fold increase in mRNA expression loss (Fig. 4C) and inflammatory score (Fig. in Nod22939iC macrophages relative to WT macrophages, which was given an arbitrary level of 1.0 T 4D, fig. S10) between the genotypes were for each gene. Results are means SEM of three independent experiments. (D) Elevated IL-1b 2939iC 2939iC abolished. secretion in MDP-stimulated Nod2 macrophages. WT and Nod2 (m/m) BMDMs were 2939iC stimulated as indicated. After 24 hours, culture supernatants were collected and secreted cytokines By contrast to the Nod2 mutation, were measured. (E) MDP induces IL-1b release by Nod22939iC (m/m) BMDMs. Macrophages were deletion of Ikkb in hematopoietic and mye- treated with MDP or LPS for 24 hours. Culture supernatants were collected and analyzed by loid cells reduced the inflammatory response immunoblotting with antibodies against IL-1b and TNFa.

736 4 FEBRUARY 2005 VOL 307 SCIENCE www.sciencemag.org R EPORTS as coincubation of macrophages with MDP plus which did not show increased inflam- many proinflammatory genes coding for cy- a TLR2 agonist (PGN) did not reduce the re- mation (11). The gain-of-function hypoth- tokines and chemokines, including IL-1b, sponse to PGN (Fig. 2D). The inhibitory esis is consistent with clinical observations TNFa, and IL-6 (22, 23), and may therefore function hypothesis is also inconsistent with made in CD patients (8, 21). be an important pathogenic factor in CD (8). in vivo findings in Nod2 knockout mice, The NF-kB signaling pathway induces Although increased transcription of many

Fig. 3. Enhanced NF-kB activation and inflammation in DSS-treated Nod22939iC mice. (A) Increased body weight loss in DSS- exposed Nod22939iC mice. Mice of either genotype were given 3% DSS in drinking water for 6 days and weighed daily. Data are means T SEM. Significant differences, *P G 0.05. (B) Typical colon appearance (upper panels) and histology (bottom panels) 11 days after initiation of DSS administration. Nod22939iC mice exhibit more inflammation and ulceration. Arrowheads, borders of ulcers. Mag- nification, 100.(C) Induction of inflammation- associated genes in colons of DSS-treated mice. Colonic RNA isolated 11 days after initiation of DSS treatment was analyzed by real-time PCR. Results are Colon sections prepared 11 days after initiation of DSS treatment were means T SEM of fold increase in normalized (relative to glyceraldehyde-3- analyzed by indirect immunoperoxidase staining for IL-6 and Cox-2. phosphate dehydrogenase mRNA) mRNA amounts in DSS-treated mice Magnification, 100.(F) Colonic NF-kB and IKK activities. Nuclear and over untreated mice of the same genotype (n 0 4pergroup).(D) cytosolic extracts of colonic mucosa prepared 0 and 11 days after initiation Elevated IL-1b and IL-6 in colons of DSS-treated Nod22939iC mice. The of DSS administration were analyzed for NF-kB DNA binding and IKK kinase indicated cytokines were measured in colonic extracts prepared 0 or (KA) activities. Protein recovery in nuclear extracts was determined by 11 days after DSS exposure. Results are means T SD (n 0 4 to 8). Significant immunoblotting with antibody against histone deacetylase (HDAC), and IK difference, *P G 0.05. (E) Immunohistochemical detection of IL-6 and Cox-2. recovery was determined with antibody against IKK (IKKa).

www.sciencemag.org SCIENCE VOL 307 4 FEBRUARY 2005 737 R EPORTS Fig. 4. IL-1b is an im- portant contributor to elevated colonic inflam- mation in Nod22939iC mice. (A and B) Increased macrophage apoptosis in Nod22939iC (m/m) mice treated with DSS. Tissue specimens prepared 0 and 11 days after initia- tion of DSS administra- tion were analyzed by TUNEL staining (A) or by TUNEL (green) plus im- munoperoxidase stain- ing for F4/80 (red) (B). Magnification: (A), 200; (B), 400.(C) Increased body weight loss in DSS- exposed Nod22939iC mice is IL-1b dependent. Mice of either genotype were given 3% DSS for 6 days with or without concom- itant treatment with IL- 1RA (100 mg/kg per day). Mice were weighed daily. Data are means T SEM. Asterisks indicate significant differences (WTversusWTIL-1-RA, m/m versus m/m IL-1-RA; P G 0.05). (D) Histological inflammation and tissue damage scores were de- termined 11 days after initiation of DSS treatment in the mice from (C). Results are means T SEM. Significant differences, *P G 0.05.

NF-kB targets was observed, the results with be mediated through the N-terminal CARD 15. Y. Weinrauch, A. Zychlinsky, Annu. Rev. Microbiol. IL-1b were unique, as it was the only pro- domains of NOD2, which may directly in- 53, 155 (1999). 16. R. A. Le Feuvre, D. Brough, Y. Iwakura, K. Takeda, N. J. inflammatory cytokine whose secretion in teract with caspase-1 or upstream caspases. Rothwell, J. Biol. Chem. 277, 3210 (2002). response to MDP was markedly elevated in Given the importance of IL-1b for the pa- 17. N. A. Thornberry, Y. Lazebnik, Science 281, 1312 Nod22939iC macrophages relative to WT thology of DSS-induced colitis in Nod22939iC (1998). 18. F. Martinon, J. Tschopp, Cell 117, 561 (2004). counterparts. Our results suggest that IL-1b mice and the imbalance between IL-1b and 19. F. R. Greten et al., Cell 118, 285 (2004). is indeed an important contributor to the IL-1-RA in CD patients (2), it would be of 20. T. Watanabe, A. Kitani, P. J. Murray, W. Strober, increased colonic inflammation in Nod22939iC interest to critically evaluate its role in CD Nature Immunol. 5, 800 (2004). 21. B. E. Sands et al., N. Engl. J. Med. 350, 876 (2004). mice, as previously suggested for CD pa- pathogenesis. 22. P. J. Barnes, M. Karin, N.Engl.J.Med.336, 1066 tients (2). (1997). Although NF-kB was thought to be the 23. L.-W. Chen et al., Nature Med. 9, 575 (2003). major effector for NOD2, it should be noted References and Notes 24. S. Rakoff-Nahoum, J. Paglino, F. Eslami-Varzaneh, 1. S. E. Girardin, J. P. Hugot, P. J. Sansonetti, Trends S.Edberg,R.Medzhitov,Cell 118, 229 (2004). that NF-kB is more effectively activated by Immunol. 24, 652 (2003). 25. Research was supported by grants from the NIH bacterial products through TLRs (see Fig. 2). 2. C. Fiocchi, Gastroenterology 115, 182 (1998). (AI43477 to M.K.; DK35108 to M.K., M.F.K., and L.E.; Thus NF-kB activation is not unique to 3. Y. Ogura et al., Nature 411, 603 (2001). and AI56075 to L.E.) and the Crohn’s and Colitis 4. J. P. Hugot et al., Nature 411, 599 (2001). Foundation of America (L.E.). S.M., L.-C.H., and L.A.B. NOD2, and its loss may not compromise 5. Y. Ogura et al., J. Biol. Chem. 276, 4812 (2001). were supported by postdoctoral fellowships from NF-kB signaling in response to bacterial 6. N. Inohara et al., J. Biol. Chem. 278, 5509 (2003). the Japan Society for the Promotion of Science, the infection. Recently, TLR signaling and a cer- 7. S. E. Girardin et al., J. Biol. Chem. 278, 8869 Cancer Research Institute, and NIH training grant (DK07202), respectively. M.K. is an American Cancer tain amount of enteric bacteria were shown (2003). 8. D. K. Podolsky, N. Engl. J. Med. 347, 417 (2002). Society research professor. to be critical for maintenance of the in- 9. C. Wahl, S. Liptay, G. Adler, R. M. Schmid, J. Clin. testinal barrier function (24), a function that Invest. 101, 1163 (1998). Supporting Online Material was suggested to deteriorate in CD pa- 10. E. Kopp, R. Medzhitov, Curr. Opin. Immunol. 15,396 www.sciencemag.org/cgi/content/full/307/5710/734/ (2003). DC1 tients (2). However, maintenance of barrier 11. A. L. Pauleau, P. J. Murray, Mol. Cell. Biol. 23,7531 Materials and Methods function is unlikely to involve NOD2. By (2003). Figs. S1 to S11 contrast, a unique function of NOD2, not 12. S. Maeda et al., unpublished observation. References and Notes 13. W. Strober, I. J. Fuss, R. S. Blumberg, Annu. Rev. provided by TLRs, is induction of IL-1b Immunol. 20, 495 (2002). 5 August 2004; accepted 8 December 2004 processing and release. This function may 14. H. C. Rath et al., Infect. Immun. 69, 2277 (2001). 10.1126/science.1103685

738 4 FEBRUARY 2005 VOL 307 SCIENCE www.sciencemag.org R EPORTS lane 4). This interaction is direct: A purified, The Kaposin B Protein of KSHV Escherichia coli–derived fusion protein of glu- tathione S-transferase (GST) and kaposin B, Activates the p38/MK2 Pathway but not GST alone, efficiently binds to purified recombinant MK2 (Fig. 1C). Parallel exper- iments with GST-DR1 and GST-DR2 proteins and Stabilizes Cytokine mRNAs mapped the interaction domain to the DR2 Craig McCormick and Don Ganem* region (Fig. 1C). Using truncated MK2 pro- teins, we also mapped the domains of MK2 Cytokine production plays a critical role in diseases caused by Kaposi’s required for interaction with kaposin B. Kapo- sarcoma–associated herpesvirus (KSHV). Here we show that a latent KSHV sin B binds to a region in the C-lobe of the gene product, kaposin B, increases the expression of cytokines by blocking the kinase domain of MK2, with residues 200 degradation of their messenger RNAs (mRNAs). Cytokine transcripts are to 270 apparently critical for the interaction normally unstable because they contain AU-rich elements (AREs) in their 3¶ (Fig. 1D) (fig. S1). This region contains a noncoding regions that target them for degradation. Kaposin B reverses this highly conserved activation segment, includ- instability by binding to and activating the kinase MK2, a target of the p38 ing a site for phosphorylation by p38 MAPK mitogen-activated protein kinase signaling pathway and a known inhibitor of (Thr222) and a binding site for the autoin- ARE-mRNA decay. These findings define an important mechanism linking hibitory C-terminal domain of MK2 (16, 17). latent KSHV infection to cytokine production, and also illustrate a distinctive These in vitro binding findings are fur- mode by which viruses can selectively modulate mRNA turnover. ther corroborated by studies that examine the colocalization of the two proteins un- Kaposi_s sarcoma–associated herpesvirus To identify proteins that interact with kapo- der various conditions in vivo. Expression (KSHV) is a g-2 herpesvirus etiologically linked sinB,weusedtheDR2andDR1repeatsto of kaposin B in cells reveals that the pro- to Kaposi_s sarcoma (KS) (1), an unusual tumor screen a library of infected cell cDNAs in tein is colocalized with MK2 in the nucleus composed of proliferating, spindle-shaped en- the yeast two-hybrid system (14). This yielded (Fig. 1E), despite the absence of a classi- dothelial cells, slit-like neovascular spaces, a clone consisting of amino acids 188 to cal nuclear localization signal in kaposin and inflammatory cell infiltration (2). KS spin- 400 of MK2 Emitogen-activated protein kinase B. MK2 is known to be exported from the dle cells, which are latently infected with (MAPK)–associated protein kinase 2^,apro- nucleus in response to inflammatory sig- KSHV, elaborate a variety of pathogenetically tein kinase whose activity is controlled by nals that activate the p38 MAPK pathway important proinflammatory cytokines and an- phosphorylation by p38, a MAPK activated by Ee.g., tumor necrosis factor–a (TNF-a) and giogenic factors (3, 4); in turn, cultured spindle inflammatory and other signals (15). Immuno- lipopolysaccharide (LPS)^ (18). It is note- cells require cytokines Eincluding interleukin-6 precipitation of FLAG epitope–tagged kapo- worthy that treatment of cells with LPS (IL-6)^ for their survival and proliferation (5). sin B results in efficient coprecipitation of (Fig. 1E) or TNF (fig. S2) causes kaposin B KSHV also causes several rare lymphoprolif- cotransfected MK2 in 293T cells, confirming and MK2 to be coexported from nucleus to erative diseases (6, 7), and these too are asso- the interaction in vivo (Fig. 1B, bottom panel, cytosol. ciated with elevated production of cytokines, notably IL-6 (6–10). However, the mechanisms linking KSHV and host cytokine production have remained obscure. Cells latently infected by KSHV express only a handful of viral genes, including the products of the kaposin locus (11) (Fig. 1A). This locus contains a small coding region Eopen reading frame (ORF) K12^ preceded by two families of 23-nucleotide GC-rich direct repeats (termed DR1 and DR2), and is tran- scribed as a single mRNA encompassing all three components. A complex translational program generates a variety of proteins from this mRNA (12). Kaposin B, the subject of this report, results from translation of the re- peats alone and consists of a series of tan- demly repeated copies of 23–amino acid peptides derived from translation of the DR2 Fig. 1. (A) Schematic map of the K12 (kaposin) locus derived from a pulmonary KS isolate of KSHV (HPRNPARRTPGTRRGAPQEPGAA) and (12). (B) Coimmunoprecipitation of kaposin B and MK2. 293T cells were cotransfected with ex- DR1 (PGTWCPPPREPGALLPGNLVPSS) re- pression vectors encoding MK2 and FLAG-tagged kaposin B. Cell lysates were immunoblotted with polyclonal antibody to MK2 (anti-MK2) (upper panel). In parallel, portions of the same lysates were peats (12, 13). Its function has heretofore immunoprecipitated with FLAG monoclonal antibody (mAb) and immunoblotted with anti-MK2 been unknown. (lower panel). (C) Recombinant full-length MK2 purified from E. coli was incubated with the indicated GST fusion proteins bound to glutathione-Sepharose beads. MK2 was detected by Western blotting with anti-MK2. (D) Kaposin B binds to a central portion of the C-lobe of MK2. Truncated Howard Hughes Medical Institute, Department of versions of MK2 were labeled with [35S]methionine by in vitro translation and tested for binding to Microbiology and Immunology, and Department of GST–kaposin B (see fig. S1 for details). NLS, nuclear localization signal; NES, nuclear export signal. (E) Medicine, University of California, San Francisco, CA Coexport of kaposin B and MK2 in response to stress signals. SLK endothelial cells were co- 94143, USA. transfected with plasmids encoding kaposin B and green fluorescent protein–MK2 fusion protein. *To whom correspondence should be addressed. After 30 hours, cells were treated with LPS (1 mg/ml) for 30 min, then fixed and stained with kaposin E-mail: [email protected] B mAb and rhodamine-conjugated secondary antibody. DAPI, 4¶,6¶-diamidino-2-phenylindole.

www.sciencemag.org SCIENCE VOL 307 4 FEBRUARY 2005 739 R EPORTS Binding of kaposin B to MK2 leads to ac- Fig. 2. Kaposin B stimulates MK2 kinase tivation of MK2, by several criteria. First, cells activity and blocks the degradation of expressing kaposin B displayed higher levels cytokine mRNAs. (A) HeLa cells were transfected with empty vector or kaposin of phosphorylated (activated) MK2 as judged B expression vector, lysed, and harvested. by immunoblotting (Fig. 2A). Second, relative Portions of the whole-cell lysates were to MK2 precipitated from control cells, en- immunoblotted with antibodies to MK2 dogenous MK2 immunoprecipitated from cells (lower panel) and phosphorylated MK2 expressing kaposin B was more active in phos- (upper panel). (B) Portions of the same phorylating the MK2 substrate heat shock pro- HeLa cell lysates were immunoprecipitated with anti-MK2, and kinase activity was tein 27 (Hsp27) (Fig. 2B). measured in vitro with recombinant Hsp27 Elevated MK2 kinase activity has a number as a substrate. (C) HeLa–Tet Off cells were of functional consequences, the best character- cotransfected with b-globin–based reporter ized of which is the stabilization of cytokine and test plasmids. After 30 hours, doxycycline transcripts and other mRNAs that contain AU- was added to the media to stop transcription. rich elements (AREs). AREs typically consist RNA was harvested at 0, 1, 2, and 4 hours after doxycycline addition; b-globin mRNA of multiple copies of the sequence AUUUA, 32 ¶ was detected with a P-labeled antisense usually located in the 3 untranslated region riboprobe. (D)HeLa–TetOffcellswere (3¶UTR) of the transcript. Their presence leads transfected with a b-globin–ARE expression to drastic destabilization of the mRNA in the vector and either an empty vector or kaposin B expression vector. One hour before addition of doxycycline, ground state (19–21), but this instability can be 1 mM hydrogen peroxide was added to one vector-transfected culture (top panel). Samples of RNA were abrogated by activation of MK2 (22). Accord- Northern blotted for b-globin mRNA as above. (E) Human foreskin fibroblasts were transfected with empty vector or kaposin B expression vector. After 48 hours, cell supernatants were collected and extracellular ingly, we tested the effects of kaposin B ex- GM-CSF and IL-6 levels were measured by ELISA. pression on this activity with the use of a standard assay of ARE-mediated mRNA de- cay (23) based on a globin reporter gene. Fig. Fig. 3. Latent KSHV infection blocks degrada- 2C shows that the b-globin mRNA reporter is tion of ARE-containing quite stable and is unaffected by cotransfection mRNAs. (A) HeLa–Tet of kaposin B. However, the insertion of an Off cells were trans- ARE Ehere, from the 3¶UTR of granulocyte- fected with b-globin– macrophage colony-stimulating factor (GM- ARE reporter 24 hours CSF)^ into the b-globin gene renders the before infection with KSHV. At 24 hours after chimeric mRNA susceptible to rapid degrada- infection, doxycycline tion, and cotransfection of kaposin B with the was added to the me- b-globin–ARE vector strikingly reverses this dia to stop transcription. effect. The magnitude of this stabilization is RNA was harvested at 0, similar to that induced by other, more potent, 1, 2, and 4 hours after triggers of the p38-MK2 pathway, including oxi- doxycycline addition; b- globin mRNA was de- dative stress (Fig. 2D). Correspondingly, trans- tected with a 32P-labeled fection of kaposin B into cells resulted in antisense riboprobe. (B) major augmentation of both GM-CSF and IL-6 Phosphorimager-based production, as determined by enzyme-linked im- quantitation of the results. Open circles, GM-CSF ARE þ vector; solid circles, GM-CSF ARE þ þ þ munosorbent assay (ELISA) of the culture medi- kaposin B; open squares, IL-6 ARE vector; solid squares, IL-6 ARE kaposin B. (C) Other KSHV um 48 hours after transfection (Fig. 2E) (fig. S3). latency genes do not block ARE-mediated mRNA decay. HeLa–Tet Off cells were cotransfected with the indicated b-globin–ARE reporter and test plasmid. After 30 hours, doxycycline was added To determine whether ARE-mediated and RNA was harvested and Northern blotted as above. (D) HeLa cells were transfected with mRNA decay is also impaired in KSHV la- plasmids encoding kaposin B or other latency-associated KSHV genes. At 30 hours after tency, we transfected HeLa–Tet Off cells with transfection, cells were lysed and immunoprecipitated with anti-MK2. MK2 kinase activity was a b-globin–ARE expression vector, then in- assayed with Hsp27 substrate and an antibody to phosphorylated Hsp27 (top panel). Total fected the cells with KSHV virions; this re- immunoprecipitated MK2 was measured with anti-MK2 (bottom panel). sulted in a latent KSHV infection. Doxycycline was then added to arrest transcription, and b- Correspondingly, the only latent gene product and 4C), to date we have detected only very globin mRNA levels were determined at 0, capable of activating MK2 kinase activity in weak binding of purified p38 to recombinant 1, 2, and 4 hours thereafter. Latently infected such transfectants was kaposin B (Fig. 3D). kaposin B in vitro (Fig. 4C). This suggests that cells displayed substantial prolongation of How does kaposin B expression lead to p38isrecruitedtokaposin-MK2complexes the half-life of the ARE-containing transcript MK2 activation? Although our understanding principally (though perhaps not exclusively) (Fig. 3, A and B). To establish that this effect of this process is still incomplete, important via its binding to MK2. The enhanced p38 ac- was due to kaposin B and not to other latency clues emerged when we examined the state of tivity observed in kaposin B–transfected cells proteins, we tested the known latent proteins activation of the other components of the p38 is an important contributor to the net state of for their ability to block ARE-mediated mRNA MAPK pathway. HeLa cells transfected with MK2 activation in such cells, because block- decay in transfected HeLa–Tet Off cells. kaposin B display activation of p38 activity as ade of p38 activity with the selective inhibitor None of the other known latency genes—those well as that of MK2 (Fig. 4A). In fact, both SB203580 results in substantial (although not encoding latency-associated nuclear antigen p38 and MKK6, an upstream kinase impli- complete) reduction of MK2 activation (Fig. (LANA) (orf73), v–cyclin D (orf72), v-FLIP cated in p38 activation, are coimmunoprecip- 4D) and cytokine release (fig. S4). (orf71), kaposin A (K12), or interferon reg- itated from cell lysates along with kaposin These results are consistent with a model ulatory factor v-IRF3—were active in ARE- B–MK2 complexes (Fig. 4B). Although MK2 (Fig. 4E) in which initial MK2 activation after dependent mRNA stabilization (Fig. 3C). binds very efficiently to kaposin B (Figs. 1C its binding by kaposin B is amplified by p38

740 4 FEBRUARY 2005 VOL 307 SCIENCE www.sciencemag.org R EPORTS Fig. 4. p38participatesinkaposinB–MK2signaling complexes. (A) HeLa cells were transfected with empty vector or kaposin B expression vector for 30 hours, lysed, and harvested. Portions of the whole-cell lysates were immuno- blotted with antibodies to p38 (middle panel) and dual- phosphorylated p38 (upper panel). Dual-phosphorylated p38 was immunoprecipitated from these lysates, and p38 activity was measured with the use of activating transcription factor 2 (ATF2) as a substrate and immunoblotting with antibody to phosphorylated ATF2 (lower panel). (B) MKK6 and p38 coimmunoprecipitate with kaposin B–MK2 complexes. 293T cells were transfected with the indicated combinations of hemagglutinin (HA) epitope–tagged kaposin B, MK2, p38, and MKK6 for 48 hours, lysed, and immunoprecipitated with HA mAb. Western blots of the immunoprecipitated material and the whole-cell lysates were probed with polyclonal antibodies to MKK6, p38, MK2, and HA. (C) p38 and MK2 proteins (purified from E. coli) were incubated overnight at 4-C with the indicated GST-fusion proteins bound to glutathione-Sepharose beads, washed, electrophoresed, and immunoblotted with specific polyclonal antibodies. (D) Inhibition of p38 partially blocks kaposin B–mediated increases in MK2 activity. The selective p38 inhibitor SB203580 was added to kaposin B and empty vector transfected HeLa cells for 1 hour, cells were lysed, and whole-cell lysates were immunoblotted with the indicated antibodies. p38 MAPK. Phosphorylation of MK2 target proteins results in a blockade Portions of these lysates were immunoprecipitated with anti-MK2 and in ARE-mediated mRNA degradation, leading to enhanced production of assayed for MK2 kinase activity. (E) Proposed model of MK2 activation proteins from ARE-containing transcripts. These proteins include by kaposin B. The reiterated DR2 repeats of kaposin B may bind multiple cytokines as well as signaling molecules such as MKK6, which in turn MK2 proteins in the nucleus, allowing for efficient phosphorylation by could amplify p38 MAPK activation. activation, leading to further activation of MK2. 9. H. G. Drexler, C. C. Uphoff, G. Gaidano, A. Carbone, 20. A. B. Shyu, M. E. Greenberg, J. G. Belasco, Genes Dev. Much remains to be learned about the mecha- Leukemia 12, 1507 (1998). 3, 60 (1989). 10. N. Nishimoto et al., Blood 95, 56 (2000). 21. G. Shaw, R. Kamen, Cell 46, 659 (1986). nisms of MK2 and p38 activation by kaposin 11. K. A. Staskus et al., J. Virol. 71, 715 (1997). 22. R. Winzen et al., EMBO J. 18, 4969 (1999). B. Binding of MK2 by kaposin B could lead 12. R. Sadler et al., J. Virol. 73, 5722 (1999). 23. C. Y. Chen, N. Xu, A. B. Shyu, Mol. Cell. Biol. 15, directly to its activation through an induced 13. Single-letter abbreviations for amino acid residues: A, 5777 (1995). Ala; C, Cys; D, Asp; E, Glu; F, Phe; G, Gly; H, His; I, Ile; 24. Dedicated to the memory of Robert Sadler, Ph.D., conformational change; we note that the re- K, Lys; L, Leu; M, Met; N, Asn; P, Pro; Q, Gln; R, Arg; S, who discovered kaposin B as a postdoctoral fellow gion on MK2 to which kaposin B binds (Fig. Ser; T, Thr; V, Val; W, Trp; Y, Tyr. but died tragically shortly thereafter. 1D) is consistent with activation via displace- 14. S. Fields, O. Song, Nature 340, 245 (1989). 15. D. Stokoe, K. Engel, D. G. Campbell, P. Cohen, M. Supporting Online Material ment of the inhibitory C-terminal autoregula- Gaestel, FEBS Lett. 313, 307 (1992). www.sciencemag.org/cgi/content/full/307/5710/739/ tory domain of MK2. Bound kaposin B could 16. W. Meng et al., J. Biol. Chem. 277, 37401 (2002). DC1 also shield activated MK2 from the action of 17. K. W. Underwood et al., Structure 11, 627 (2003). Materials and Methods 18. K. Engel, A. Kotlyarov, M. Gaestel, EMBO J. 17, 3363 Figs. S1 to S4 cellular phosphatases. As to how p38 becomes (1998). activated, because cytokines such as IL-6 can 19. C. Y. Chen, A. B. Shyu, Trends Biochem. Sci. 20,465 28 September 2004; accepted 15 December 2004 activate the p38 pathway, elevated cytokine (1995). 10.1126/science.1105779 release from kaposin B–expressing cells may promote an autocrine or paracrine amplifica- tion loop that further enhances p38 activity. Mutualistic Fungi Control Crop Irrespective of its mechanistic details, the finding that kaposin B activates the p38-MK2 Diversity in Fungus-Growing Ants pathway forges an important biochemical link between KSHV infection and the enhanced Michael Poulsen* and Jacobus J. Boomsma cytokine production that characterizes so many of its associated disease states, and provides Leaf-cutting ants rear clonal fungi for food and transmit the fungi from a striking example of virus-mediated modu- mother to daughter colonies so that symbiont mixing and conflict, which lation of mRNA stability. result from competition between genetically different clones, are avoided. Here we show that despite millions of years of predominantly vertical References and Notes transmission, the domesticated fungi actively reject mycelial fragments from 1. Y. Chang et al., Science 266, 1865 (1994). neighboring colonies, and that the strength of these reactions are in 2. B. Ensoli, M. Sturzl, Cytokine Growth Factor Rev. 9,63 proportion to the overall genetic difference between these symbionts. Fungal (1998). incompatibility compounds remain intact during ant digestion, so that fecal 3. B. Ensoli et al., Science 243, 223 (1989). 4. B. Ensoli, G. Barillari, R. C. Gallo, Immunol. Rev. 127, droplets, which are used for manuring newly grown fungus, elicit similar 147 (1992). hostile reactions when applied to symbionts from other colonies. Symbiont 5. S. A. Miles et al., Proc. Natl. Acad. Sci. U.S.A. 87, control over new mycelial growth by manurial imprinting prevents the rearing 4068 (1990). 6. K. V. Komanduri, J. A. Luce, M. S. McGrath, B. G. of multiple crops in fungus gardens belonging to the same colony. Herndier, V. L. Ng, J. Acquir. Immune Defic. Syndr. Hum. Retrovirol. 13, 215 (1996). Ant fungiculture arose in South America growing (attine) ants (1). All extant attine 7. H. Asou et al., Blood 91, 2475 (1998). È 8. H. G. Drexler, C. Meyer, G. Gaidano, A. Carbone, about 50 to 60 million years ago in the ants ( 210 described species in 13 genera) Leukemia 13, 634 (1999). ancestor of the New World tribe of fungus- are obligately dependent on this symbiosis,

www.sciencemag.org SCIENCE VOL 307 4 FEBRUARY 2005 741 R EPORTS which mostly involves Lepiotaceaeous fungi identical banding patterns within fungus species (1.40 for A. echinatior and 1.54 for (order Agaricales, division Basidiomycota) gardens (21). A. octospinosus) were less hostile than (1–3). The symbionts of many lower attine Mycelial incompatibility was highly var- between species (1.98). This result corre- ants are closely related to free-living fungi iable among pairs of clones (Fig. 1B) and sponded to lower relative genetic distances (4), but those of Atta and Acromyrmex leaf- was positively correlated with genetic dis- between fungi within (0.20 for A. echinatior cutting ants have unique coevolved adapta- tance (Mantel test: r 0 0.855, P G 0.0001) and 0.23 for A. octospinosus) than between tions as a specialized crop (5). The ants (Fig. 1C). Average reactions within ant ant species (0.29). An effect likelihood ratio provide the fungus with fresh substrate and protection against competitors and pathogens (6–8), and virgin ant queens carry their mother_s symbiont when leaving their colo- ny to mate and disperse (9). This vertical transmission of the symbiont is expected to stabilize the mutualism by aligning the re- productive interests of the partners (10–12). However, horizontal exchange of symbionts may occur during cofounding of colonies or symbiont theft after garden loss (5, 13–16). If hostile interactions between sym- bionts reduce productivity, the introduction of an alien fungus clone will harm not only the resident fungus but also the ant hosts Fig. 1. (A)AnA. octospinosus fungus garden and (B) typical examples of the four degrees of mycelial incompatibility between fungus clones in vitro. 0 shows fully compatible clones with no demarcation (10, 11, 16). Ants in full control could pos- zone; 1 shows weakly incompatible with a slight, but clearly visible, demarcation zone; 2 shows sibly avoid these costs by rearing genetically incompatible clones with a distinct demarcation zone; and 3 shows strongly incompatible clones with a different symbionts in separate nest cham- broad demarcation zone and brown coloration of mycelium and/or medium. (C) The relation between bers, but previous studies have suggested pairwise degree of incompatibility and genetic distance (171 different combinations, each replicated six that colonies rear only a single fungus clone times) (fig. S2). The area of each circle is proportional to the number of replicate combinations producing (5, 16–18). This implies that the ants do not a given result. Controls are not shown, becauses they were all fully compatible (21). have agricultural practices of exchanging crops, nor do they have multiple crops, which are developments that have been essential in human agriculture (19). Here we show that single-crop ant farming is actively imposed by the fungal symbiont. We used fungus gardens (Fig. 1A) that were cultivated by 18 colonies of two sym- patric species of Panamanian leaf-cutting ants, Acromyrmex echinatior and A. octospinosus. These two ant species have never been observed to hybridize, but their clonal fungi belong to the same genetically diverse (fig. S1) (16, 20, 21). Mycelial incompat- ibility between fungi from different colo- nies was assessed by inoculating pairs of fungus that were grown 1.5 cm apart on potato dextrose agar (PDA) medium (fig. S2) (21). After 2 months, mycelial compatibility could be scored on a scale from 0 to 3 (Fig. 1B) (21, 22). We also obtained genetic fingerprints of the fungi by using amplified fragment length polymorphism (AFLP) to estimate relative genetic distance (percent- age of bands not homologous) between pairs of clones and to confirm that each source colony cultivated a single clone (21). The Fig. 2. (A)AnA. echinatior fungus garden with a fecal droplet (encircled) [photo courtesy of Klaus latter was done by AFLP analysis of five to Lechner]. (B) Typical examples of the categories of reaction of plated fungi (after 24 hours) (21)toward fecal droplets produced by workers from other colonies of A. echinatior or A. octospinosus from the same six independently isolated samples of the site. 0 shows complete absorption indicating full compatibility; 1 shows partial absorption; 2 shows same fungus garden from each of 10 col- distinct mycelium growth on the droplet but no absorption; 3 shows no mycelial growth on the droplet onies. Alignment of these profiles showed or in the direct vicinity of the droplet; 4 shows active avoidance of the droplet by fungal hyphae, with droplet turning light to dark brown; and 5 shows complete rejection of the droplet and very dark coloration of both droplet and surrounding mycelium. (C) The pairwise degree of incompatibility Institute of Biology, Department of Population Biology, between fungal fragments maintained by ants and fecal droplets after 24 hours, plotted against genetic , Universitetsparken 15, DK- distance (garden fragments and fecal droplets from eight different colonies were combined in all possible 2100 Copenhagen, Denmark. ways and replicated eight times). Circle area is proportional to number of combinations producing a *To whom correspondence should be addressed. given result and controls are not shown [67.9% showed full compatibility (0); 30.4% showed partial E-mail: [email protected] absorption(1);and1.7%showeddistinctmyceliumgrowthonthedroplet,butnoabsorption(2)](21).

742 4 FEBRUARY 2005 VOL 307 SCIENCE www.sciencemag.org R EPORTS (ELR) test confirmed that incompatibility is P G 0.0001) (Fig. 2C). ELR testing showed derived fecal droplets (Figs. 2 and 4), and the related to fungal genetic distance (c2 0 2569, that the magnitude of this effect was not hostile ant behavior toward alien droplets degrees of freedom (df) 0 1, P G 0.0001) and due to ant species combination (c2 0 1.357, (Fig. 3) and alien fungus fragments (16), not an effect of ant species combination (c2 0 df 0 2, P 0 0.5074) but only to the genetic show that the host ants and their resident 2.199, df 0 2, P 0 0.3330) (21). distance of symbionts (c2 0 432.4, df 0 1, symbiont jointly prevent the introduction of Acromyrmex worker ants actively discrim- P G 0.0001) (21). competing fungus clones, which agrees with inate between fragments of their resident In the same experimental colonies, there evolutionary theory on host symbiont con- fungus and genetically different symbionts was a significant association between the flict over symbiont mixing (10, 11). In- (16), which confirms predictions that the ants likelihood that ants would remove alien fecal compatibility reactions (brown coloration of have a strong short-term interest in maintain- droplets and the genetic distance between the mycelium) have also been observed in ing genetically pure gardens (11). The ants resident and the droplet-producing fungus interactions between the ants_ fungal sym- reject alien fungus fragments for at least 1 (Mantel test: r 0 0.721, P G 0.0001) (Fig. bionts and pathogenic fungi (26, 27) and week after being deprived of their resident 3A) and the droplet-fungus incompatibility probably facilitate the detection of fungal garden, which indicates that avoidance of reaction (Mantel test: r 0 0.651, P G 0.0001) disease. However, the present results suggest symbiont competition, at least initially, over- (Fig. 3B). Weeding behavior around the that these recognition abilities have been rides symbiont replacement behavior, which droplets, which resulted in the accumulation enhanced after fungus domestication, partic- is in their long-term interest (16). This sug- of additional fungal waste material (8, 21), ularly in the higher attine ants (3, 5). Direct gests that the weeding behavior of worker was associated with the same predictor assessments by the ants (16, 28, 29) and the ants may be directly linked to fungal in- variables EMantel tests: r 0 0.700, P G derived recognition mechanism by the fecal compatibility compounds. 0.0001 (Fig. 3C) and r 0 0.686, P G 0.0001 droplets (G24 hours; Figs. 2 and 4) are much All fungus-growing ants manure newly (Fig. 3D)^. Droplet removal and garbage ac- faster than recognition through mycelial grown mycelium with their own feces cumulation were likewise correlated (Mantel contact, which takes 5.6 days on average (23, 24). In this process, fungal enzymes, test: r 0 0.728, P G 0.0001) (21). This (range, 3 to 7 days), even if tufts of alien which help break down plant material, pass suggests that the ants actively discard alien mycelium are applied directly on mats of through the ant gut into fecal droplets (25), fecal droplets and any mycelium that has plated symbionts (Fig. 1B) (21). The chem- which are deposited on fresh leaves (Fig. 2A) been in touch with these droplets. ical nature of the incompatibility compounds or directly on the fungus garden, where they In a final experiment, we found that remains unknown, but they could be similar are readily absorbed (2). If incompatibility initially incompatible interactions between to those of free-living basidiomycetes or be compounds remained unaffected by ant diges- fecal droplets and fungi became compatible derivates of evolutionary-derived enzymes of tion, we would expect that fecal droplets when ants were forced to feed on an the symbionts (30). secondarily might have become defensive incompatible alien symbiont for 10 days. Neither direct recognition of alien symbi- extended phenotypes of the resident symbiont. The ants_ new fecal droplets also became onts by the ants nor mycelial incompatibility We tested this idea by examining the reactions incompatible with their original resident can explain why Acromyrmex leaf-cutting of fungus and ants toward fecal droplets from fungus (Fig. 4, B and C) (21). An ELR test ants are restricted to rear a single symbiont other colonies when the droplets were applied showed a significant main effect of the ant clone per colony. Abundant genetic variation to fungus garden fragments of live experimen- fungus combination (c2 0 9.498, df 0 1, P 0 between clones is available, and mature nests tal colonies (fig. S3) (21). 0.0021) and a significant interaction between of A. octospinosus and A. echinatior are Incompatibility reactions of fungi toward the ant fungus combination and time (c2 0 usually compartmentalized with multiple fecal droplets from nonresident ants were 11.58, df 0 1, P 0 0.0007) (21). Controls did gardens that could keep symbiont strains scored on a scale ranging from 0 to 5 after 24 not show significant changes in compatibil- separated. The present study suggests that hours (Fig. 2B). There was a significant ity, which confirmed that the compounds association between the intensity of the responsible for incompatibility reactions are reactions and the genetic distance between exclusively fungus-derived. the resident fungus and the fecal droplet– The parallel reactions of resident fungi producing fungus (Mantel test: r 0 0.960, toward alien mycelia (Fig. 1) and fungus-

Fig. 3. (A and B) Fecal droplet removal and (C and D)amountofgarbage (milligrams of fresh weight) deposited 24 hours after application of fecal droplets that were obtained from the same eight-by-eight combinations of fungus gardens and ants used in Fig. 2 (21). Data are plotted against genetic distance between the resident and the droplet-producing fungus [(A) and (C)] and the degree of fecal-droplet in- compatibility (0 to 5) [(B) and (D)]. Fig. 4. Fungus incompatibility reactions toward Circle area is proportional to the resident and foreign fecal droplets, before and number of combinations producing a after force-feeding the ants that produced the given result. Controls are not plotted, droplets for 10 days. Two colonies (Ao165 because resident fecal droplets were and Ae209), which were initially droplet- never removed and the average amount incompatible (Fig. 3), were used for reciprocal of garbage deposited was 1 T 0.3 mg force-feeding treatments (B and C)and (mean T SE), which was substantially controls (A and D). Mean reactions T SE (n 0 less than the mean garbage accumula- 3 replicate pairings) displayed by fungus Ao165 tion of all the test combinations (4 T 0.4 mg) (21). (squares) and Ae209 (circles) are plotted.

www.sciencemag.org SCIENCE VOL 307 4 FEBRUARY 2005 743 R EPORTS the ants_ manuring practice is the decisive References and Notes 22. E. M. Hansen, J. Stenlid, M. Johansson, Mycol. Res. factor that constrains colonies to rearing a 1. I. H. Chapela, S. A. Rehner, T. R. Schultz, U. G. 97, 1229 (1993). Mueller, Science 266, 1691 (1994). 23. M. M. Martin, Science 169, 15 (1970). single clone of symbiont. Obligate manuring 2. N. A. Weber, Ed., Gardening Ants: The Attines 24. T. Murakami, S. Higashi, J. Ethol. 15, 17 (1997). with ant feces allows the resident fungus to (American Philosophical Society, , 1972). 25. S. Rønhede, J. J. Boomsma, S. Rosendahl, Mycol. Res. control the genetic identity of new gardens in 3. T. R. Schultz, U. G. Mueller, C. R. Currie, S. A. Rehner, 108, 101 (2004). in Ecological and Evolutionary Advances in Insect- 26. C. R. Currie, personal communication. the nest, causing the removal of unrelated Fungal Associations, F. Vega, M. Blackwell, Eds. 27. M. Poulsen, personal observation. fungi before they contribute to ant feeding (Oxford Univ. Press, Oxford, 2004), pp. 149–190. 28. A. M. M. Viana et al., Chemoecology 11, 29 (2001). and the production of compatible fecal 4. P. Villesen, U. G. Mueller, T. R. Schultz, R. M. M. 29. U. G. Mueller, J. Poulin, R. M. M. Adams, Behav. Ecol. Adams, A. C. Bouck, Evolution 58, 2252 (2004). 15, 357 (2004). droplets (Fig. 4). The manurial imprint of 5. U. G. Mueller, S. A. Rehner, T. R. Schultz, Science 281, 30. J. J. Worrall, Mycologia 89, 24 (1997). the fungus therefore makes ant agriculture 2034 (1998). 31. P. Ridley, P. E. Howse, C. W. Jackson, Experentia dependent on a single symbiont that is very 6. M. Bass, J. M. Cherrett, Ecol. Entomol. 19, 215 (1994). (Basel) 52, 631 (1996). 7. R. D. North, C. W. Jackson, P. E. Howse, Trends Ecol. 32. U. G. Mueller, Am. Nat. 160 (suppl., 67 (2002). difficult to replace and impossible to com- Evol. 12, 386 (1997). 33. D. K. Aanen et al., Proc. Natl. Acad. Sci. U.S.A. 99, bine with secondarily acquired symbionts. 8. C. R. Currie, A. Stuart, Proc. R. Soc. London Ser. B 14887 (2002). The manurial imprint of fecal droplets is 268, 1033 (2001). 34. H. Katoh, T. Miura, K. Maekawa, N. Shinzato, T. 9. B. Ho¨lldobler, E. O. Wilson, The Ants (Springer Verlag, Matsumoto, Mol. Ecol. 11, 1565 (2002). a clear example of fungal signaling affecting Berlin, 1990). 35. J. Diamond, Science 281, 1974 (1998). ant behavior and is possibly more important 10. S. A. Frank, Proc.R.Soc.LondonSer.B263, 339 (1996). 36. We thank B. E. N. Markussen, S. M. Mathiasen, and than suggested fungal signaling for directing 11. S. A. Frank, Evolution 57, 693 (2003). S. Rønhede for laboratory assistance; D. R. Nash and the choice of food plants by workers to avoid 12. E. A. Herre, N. Knowlton, U. G. Mueller, S. A. Rehner, J. S. Pedersen for detailed statistical advice; D. K. Aanen, Trends Ecol. Evol. 14, 49 (1999). S.A.O.Armitage,M.J.Cafaro,C.R.Currie,D.R.Nash, an overdose of toxins (31, 32). Similar 13. S. W. Rissing, G. B. Pollock, M. R. Higgins, R. H. J. S. Pedersen, S. Taerum, and J. M. Thomas for fruitful manipulative effects of resident fungi toward Hagen, D. R. Smith, Nature 338, 420 (1989). discussions and comments; the Smithsonian Tropical competing clones may have convergently 14. D. Bekkevold, J. Frydenberg, J. J. Boomsma, Behav. Research Institute for providing facilities to work in Ecol. Sociobiol. 46, 103 (1999). Panama; and the Autoridad Nacional del Ambiente y el evolved in the fungus-growing termites, 15. R. M. M. Adams, U. G. Mueller, A. K. Holloway, A. M. Mar for sampling and export permissions. Supported by where the fungus substrate is entirely derived Green, J. Narozniak, Naturwissenschaften 87,491 grants from the Danish Natural Science Research from fecal matter (33). This could explain (2000). Council and the Carlsberg foundation (J.J.B.) and a 16. A. N. M. Bot, S. A. Rehner, J. J. Boomsma, Evolution Ph.D. stipend from the University of Copenhagen (M.P.). why fungus-growing termites also rear single 55, 1980 (2001). clones of fungus, despite initiating colonies 17. A. M. Green, U. G. Mueller, R. M. M. Adams, Mol. Supporting Online Material with horizontally acquired fungal spores Ecol. 11, 191 (2002). www.sciencemag.org/cgi/content/full/307/5710/741/ 18. U. G. Mueller, S. E. Lipari, M. G. Milgroom, Mol. Ecol. DC1 (33, 34). None of these constraints have ever 5, 119 (1996). Materials and Methods applied to human agriculture because crops 19. Y. Zhu et al., Nature 406, 718 (2000). Figs. S1 to S3 and expertise have been culturally trans- 20. T. D. Schultz, D. Bekkevold, J. J. Boomsma, Insect Soc. References 45, 457 (1998). mitted between tribes from the beginning of 21. Materials and methods are available as supporting 25 October 2004; accepted 1 December 2004 human farming (35). material on Science Online. 10.1126/science.1106688

744 4 FEBRUARY 2005 VOL 307 SCIENCE www.sciencemag.org EW RODUCTS N http://science.labvelocity.comP

Cell Cycle Assay on a fluorescence-blocking, microporous-membrane insert in a The Cell Cycle Assay makes cell cycle analysis easier for applications multiwell format.The BD BioCoat Angiogenesis System: Endothelial in immunology, cancer drug discovery, and cell health profiling. Cell Invasion is a robust cell culture insert system designed to Staining reagents are pre-combined in a single optimized format, simplify endothelial screening assays. eliminating the need for complex reagent preparation following cell BD Biosciences For information 800-343-2035 www.bdbiosciences.com fixation. The assay provides detailed protocols for use with either individual tubes or 96-well microtiter plates.The assay’s automated Hot Start Polymerase analysis capability displays results in both histogram and statistical LA Taq Hot Start DNA polymerase is designed for polymerase formats, making it a convenient method of screening and optimiz- chain reaction (PCR) amplification of long DNA fragments (up to ing lead drug candidates for their ability to induce cell cycle arrest 43 kb) with increased specificity and reduced background. LA Taq and to kill proliferating cells. Together with Guava’s Personal Cell Hot Start contains LA Taq,which provides higher fidelity than con- Analysis system and CellHealth Profiling assays, the new Cell Cycle ventional Taq DNA polymerase, plus a neutralizing monoclonal Assay provides faster, easier cell analysis capabilities at the antibody to Taq.This antibody binds to the polymerase until the researcher’s benchtop. temperature is elevated during the first denaturation step, pre- Guava Technologies For information 510-576-1427 www.guavatechnologies.com venting nonspecific amplification due to mispriming or formation of primer dimers during the reaction assembly. The PCR products Genome Bioarray generated are suitable for cloning into T vectors. The new CodeLink Mouse Whole Genome Bioarray, a new addition Takara For information 888-251-6618 www.takaramirusbio.com to the CodeLink microarray platform portfolio, is the third whole- genome bioarray from GE Healthcare this year. For researchers Electronic Pipette using the mouse as a model system for human disease study, this The Transferpette electronic pipette features an new bioarray offers the entire mouse genome on a single array for ergonomic design and an innovative tip cone gene expression profiling. Designed for comprehensive genome- that ensures low-force tip release using virtu- wide mouse gene expression profiling, this bioarray contains a set ally all standard tips. Five convenient operating of about 35,000 probes targeting more than 36,000 mouse tran- modes increase productivity. It is available in scripts on a single slide. three volume ranges for dispensing 2 µl to GE Healthcare For information +44 (0) 1494 498 068 www.gehealthcare.com 1000 µl. Brandtech Scientific For information 888-522-2726 Real-Time MultiPlex PCR www.brandtech.com The new QuantiTect MultiPlex PCR NoROX Kit provides a simple procedure for quantitative, real-time, multiplex polymerase chain Lab Refrigerator reaction (PCR) on real-time cyclers that do not use ROX dye for The Medicool Model SR-L6110W is a new undercounter laboratory fluorescence normalization. For other real-time cyclers, the Quanti- refrigerator designed to meet the temperature stability demands of Tect Multiplex PCR Kit (with ROX dye) is also available. Both kits clinical, life science, pharmaceutical, biotechnology, and industrial support the maximum multiplex capacity of the latest real-time laboratories. It offers a temperature range of 1°C to 14°C; an cyclers, and enable accurate and upward-angled, door-mounted microproces- sensitive quantification of two, sor control with LED display; an integrated three, or even four complemen- For more information visit GetInfo, automatic alarm system with audible and tary DNA or genomic DNA tar- Science's new online product index at visual notification; alarm contacts and access gets in parallel in the same tube. http://science.labvelocity.com port for auxiliary monitoring and alarms sys- As few as 10 copies of each tar- tems; and forced-air circulation for superior get sequence can be detected. From the pages of GetInfo, you can: temperature control and uniformity. Both kits make real-time, multi- • Quickly find and request free information Sanyo For information 800-858-8442 plex PCR easy through the pre- on products and services found in the pages www.sanyobiomedical.com optimized master mix and dedi- of Science. cated protocols, which have • Ask vendors to contact you with more Nitric Oxide Assay Kits been tested for use with a wide information. Nitric Oxide and Total Nitric Oxide Assay range of real-time cyclers and • Link directly to vendors' Web sites. Kits are available under the Endogen brand. sequence-specific probes. The former is a complete kit for the quanti- Qiagen For information 800-426-8157 tative determination of nitrite and nitrate in www.qiagen.com biological fluids. The latter is a complete kit for the quantitative determination of total nitric oxide in biological fluids. Endothelial Cell Invasion System Pierce Biotechnology For information 800-487-4885 www.endogen.com Endothelial cell invasion is a critical aspect of the neovascularization cascade that occurs during many normal and pathological processes Newly offered instrumentation, apparatus, and laboratory materials of interest to such as development, wound healing, and tumorigenesis.Traditional researchers in all disciplines in academic, industrial, and government organizations are angiogenesis assays are time-consuming and labor intensive, using featured in this space. Emphasis is given to purpose, chief characteristics, and availabil- ity of products and materials.Endorsement by Science or AAAS of any products or mate- microporous membranes and involving fixing, staining, and manual rials mentioned is not implied.Additional information may be obtained from the manu- counting of invaded cells. BD Biosciences has developed an easy-to- facturer or supplier by visiting www.science.labvelocity.com on the Web, where you can use, high-throughput Endothelial Cell Invasion System that is based request that the information be sent to you by e-mail, fax, mail, or telephone.

www.sciencemag.org SCIENCE VOL 307 4 FEBRUARY 2005 745 Gordon Research Conferences 2005 Summer and Fall Meetings VISIT THE frontiers of science GO TO A GORDON CONFERENCE

GRC is a nonprofit organization managed by and for the benefit of the scientific community. The 2005 Summer and Fall Gordon Research Conferences will be held in New England and Montana in the United States and international meetings will be held in France, China, Switzerland, Italy and Oxford, UK.

Conferee Adult Guest FEES: SUMMER / FALL 2005 Single Double Off-Site Single Double Off-Site Connecticut $760 $720 $575 $560 $545 $400 Massachusetts $725 $660 $575 $550 $485 $400 Maine (Colby College / Bates College) $725 $660 $575 $550 $485 $400 Maine (University of New England) $765 $765 $575 $590 $590 $400 Montana $1,130 $915 $800 $955 $740 $625 $725 $660 $575 $550 $485 $400 Rhode Island $760 $720 $575 $560 $545 $400 France (Aussois) $900 $825 $650 $725 $650 $475 DIRECTOR France (Roscoff) $875 $770 $635 $700 $595 $460 Nancy Ryan Gray France (Roscoff - Graduate Seminar) $300 $250 N/A $300 $250 N/A Hong Kong $860 N/A $615 $685 N/A $440 DIRECTOR EMERITUS Oxford (Magdalen College) $1,025 $1,025 $725 $850 $850 $550 Carlyle B. Storm Oxford (The Queen’s College) $1,000 $1,000 $880 $825 $825 $705 Switzerland $1,075 $975 $875 $900 $800 $700 DIRECTOR EMERITUS , Italy $865 $770 $530 $690 $595 $355 Alexander M. Cruickshank

The Gordon Research Conferences web site at http://www.grc.org/ contains the most up-to-date information we have for any given Confer- ence. Be sure to take a look at the scientific program or any other information that has been posted. Applications to any Conference can be submitted via our web site at http://www.grc.org/application/. A printed form may be obtained from our web site or requested from:

Gordon Research Conferences E-mail: [email protected] P.O. Box 984 FAX: 401-783-7644 West Kingston, RI 02892-0984 Phone: 401-783-4011, ext. 100

List of meetings with confirmed sessions/themes and speakers as of January, 2005 (discussion leaders are italicized):

ADVERSE DRUG REACTIONS • Pharmacogenetics • BioMEMs & MEMS CONNECTICUT COLLEGE (Michel Eichelbaum / Munir Pirmohamed / (Yoshinobu Baba / Andreas Hierlemann / NEW LONDON, CT Matthias Schwab / Leif Bertilsson) Alexandra Fuchs) JUN 5-10, 2005 • Drug-Induced Long QT Syndrome • Nanowires(Claire Darby / JACK UETRECHT, CHAIR (Peter Siegl / John Mitcheson / Borje Darpo) Donald Fitzmaurice / Reginald Penner / THOMAS KAWABATA, VICE CHAIR Bernadette Quinn) ______• Nanopores • Keynote: The Danger Hypothesis and (Li Sun / Lydia Sohn / Henry S. White) Adverse Drug Reactions • Microarrays (Polly Matzinger) (Larry Morrison / John T. McDevitt / ANALYTICAL CHEMISTRY • Mechanistic Aspects of Hans Lehrach / Robert M. Corn) THE ROSCOFF BIOLOGICAL STATION Idiosyncratic Drug Reactions • Open Session ROSCOFF, FRANCE (Kevin Park / Werner Pichler / Lance Pohl / (D. Jed Harrison) JUN 12-17, 2005 Craig Svensson) RICHARD CROOKS, CHAIR • Basic Immunology with Relevance to GORDON-KENAN GRADUATE RESEARCH D. JED HARRISON, VICE CHAIR Adverse Drug Reactions SEMINAR: ANALYTICAL CHEMISTRY (Cynthia Ju / Juan LaFaille / THE ROSCOFF BIOLOGICAL STATION • Microanalytical Systems I: Fluidics Andrew Fontenot) ROSCOFF, FRANCE (Dick Crooks / Andreas Manz / • Predictive Screens JUN 10-12, 2005 Thomas Laurell) (Tim Ryan / Bruce Car / Greg Slatter / RICHARD CROOKS, CHAIR • Microanalytical Systems II: Cells Randy Peterson) (Jörg P. Kutter / Jon M. Cooper / • Role of Regulatory Agencies in The Gordon-Kenan Graduate Research Seminar Elisabeth Verpoorte / Helene Andersson) Encouraging and Enabling Research in on Analytical Chemistry is a two-day Gordon • Microanalytical Systems III: Nanofluidics Idiosyncratic Drug Reactions Conference-style meeting exclusively for graduate (Elizabeth Zubritsky / (Kenneth Hastings / John Senior) students and postdoctoral . Speakers will be J. Michael Ramsey / Paul Bohn) • Hepatotoxicity chosen from among the attendees. Immediately • Microanalytical Systems IV: Systems (Paul Watkins / Chris Day / following the Seminar, the Gordon Research (Andrew deMello / Antonio J. Ricco / Dan Burns / James Boyer) Conference on Analytical Chemistry will take place Tom van de Goor / Takehiko Kitamori) • Selected Abstracts for Oral Presentation at the same location. (Thomas Kawabata) visit the frontiers of science: www.grc.org ANGIOGENESIS & MICROCIRCULATION • Therapeutic Applications • Protein Synthesis, Degradation & RNA SALVE REGINA UNIVERSITY (David Peritt / Patrizia Rovere-Querini) Modification NEWPORT, RI (Patrick Dennis / Dieter Soll / AUG 14-19, 2005 ______Julie Maupin-Furlow / Paola Londei / ELISABETTA DEJANA, CHAIR Arina Omer) DOUGLAS HANAHAN, VICE CHAIR • Metabolism (Richard Shand / Betina Siebers / APPLIED & ENVIRONMENTAL MICROBIOLOGY • Basic Concepts in Imke Schroeder / Mechke Pohlschroder / CONNECTICUT COLLEGE Vascular Morphogenesis John Leigh) NEW LONDON, CT (Elisabetta Dejana / Marc A. Krasnow / • Archaeal Ecology & Diversity JUL 24-29, 2005 Marc Tessier Lavigne) (Karl Stetter / Edward Delong / GERARD MUYZER, CHAIR • Blood Vessel Guidance Richard Seifert / Robert Goodman / KENNETH NEALSON, VICE CHAIR (Federico Bussolino / Christer Betzholtz / Christa Schleper / Reinhard Rachel) David Anderson / Anne Eichman) • The Need for Microbial Ecophysiology • Bone Marrow Derived Hemangiogenic (Doug Capone / Gijs Kuenen) ______Progenitors • Cultivating the Uncultured (Shahin Raffii / Eli Keshet / (Harold Drake / Slava Epstein / Stephanie Dimmeler) Karsten Zengler / Martin Hahn / ASSISTED CIRCULATION • The Development of the Cardiovascular Jared Leadbetter) BIG SKY RESORT System in Lower Vertebrates • Novel Microbial BIG SKY, MT ( / Brant Weinstein / (Svetlana Dedysh / Oded Béjà / Marc Strous) AUG 21-26, 2005 Peter Carmeliet / Didier Stainier) • Sustainable Applications of Industrial ERIC ROSE, CHAIR • Regulation of Angiogenesis by Hypoxia and Biotechnology ALAN SNYDER, VICE CHAIR Growth Factors (Claudia Schmidt-Dannert / Bernard Hauer / (Peter Carmeliet / Celeste Simon / Douglas Cameron / Lonnie Ingram / • Intro Session P.J. Ratcliffe / Lena Claesson-Welsh / Uwe Sauer) (Eric Rose / Marvin Konstam / Leslie Miller / Laura Benjamin) • Patterns in Microbial Diversity Sharon Hunt / James Long) • Growth Factors and Their Signalling (Matthew Kane / Lise Øvreas / • Present and Emerging Technologies Pathways Brendan Bohannan) (Alan Snyder / James Anderson / (Harold F. Dvorak / Kari Alitalo / • Microbial Consortia and Symbioses Robert Kormos / William Wagner / / Masabumi Shibuya / (Nicole Dubilier / Edward Ruby / Gerson Rosenberg) David Cheresh / Donald McDonald) Jörg Overmann / Shana Goffredi / • Preventing and Managing • The Influence of Microenvironment in Andreas Brune) Adverse Events (1) Vascular Morphogenesis • Ecology of Aquatic Viruses (William Holman / Mariel Jessup / Frank Lowy) (Michael Gimbrone / / (John Paul / Curtis Suttle / • Preventing and Managing Luisa Iruela Arispe / Hellmut Augustin / Markus Weinbauer / Corina Brussaard) Adverse Events (2) Raghu Kalluri) • Metagenomics in Applied and (Lynn Warner Stevenson / Ann Marie Schmidt • Paracrine Cellular Signalling Environmental Microbiology / Robert Robbins / Sam Goldhaber / (Denisa Wagner / Pat D’Amore / (Jürgen Eck / Tom Isenbarger / Gene Tyson / Diane Meier) William Sessa / Jeff Pollard) Rolf Daniel / Rachel Poretsky) • Clinical Trials • Novel Strategies to Inhibit Angiogenesis in • Special Lecture (Sean Tunis / Annetine Gelijns / In Vivo Models (Ken Nealson / ) Alan Moskowitz / Daniel Heitjan) ( / Rakesh Jain / • Challenges to Technological Innovations Gabriele Bergers / Doug Hanahan) ______(John Watson / Beverly Lorell / John Woodard / Paul Citron / ______Timothy Baldwin) • Recovery ARCHAEA: ECOLOGY, METABOLISM & (Robert Kormos / Guillermo Torre / MOLECULAR BIOLOGY APOPTOTIC CELL Christine Morevick) MAGDALEN COLLEGE RECOGNITION & CLEARANCE • Cell Transplantation OXFORD, UK CONNECTICUT COLLEGE (Silviu Itescu / Richard Weisel / AUG 14-19, 2005 NEW LONDON, CT Kenneth Chien / Katherine Verfaille / PAUL BLUM & JUN 19-24, 2005 Martin Leon) JOHN VAN DER OOST, CO-CHAIRS ROBERT SCHLEGEL & IMKE SCHROEDER & PATRICK WILLIAMSON, CO-CHAIRS MALCOLM WHITE, CO-VICE CHAIRS ______MICHAEL HENGARTNER & KODI RAVICHANDRAN, CO-VICE CHAIRS • Key Note Talk & Archaeal Virology (Karl Stetter / Mark Young / ATHEROSCLEROSIS • The PS Receptor David Prangishvili) KIMBALL UNION ACADEMY (Valerie Fadok / Peter Henson / • Evolutionary Genomics MERIDEN, NH Andreas Lengeling) (Mark Young / Eugene Koonin / James Lake / JUN 19-24, 2005 • Identification of the Apoptotic Target Patrick Forterre / Xu Peng) ISRAEL CHARO & (Robert Schlegel / Kirsten Lauber / • Functional Genomics & Systems Biology ALAN DAUGHERTY, CO-CHAIRS Giovanna Chimini / Valerian Kagan) (Mike Adams / Rolf Bernander / Greg Ferry) ELIZABETH NABEL, VICE CHAIR • Autoimmunity • Biomedicine & Biotechnology (Patrick Williamson / (John Williams / Shiladitya DasSarma / Dennis • Immune Function / Glen Matsushima) Sprott / Jonathan Trent / (Goran K. Hansson / Goran K. Hansson / • Recognition in Invertebrates Robert Kelly / Paul Eckburg) Ziad Mallat / Joseph L. Witztum / Amy Major / (Yoshinobu Nakanishi / Michael Hengartner / • RNA Synthesis & Degradation Stewart Whittman) Akiko Shiratsuchi / Loriano Ballarin) (Dieter Soll / John Reeve / Michael Thomm / • Inflammatory Processes • Diversity of Recognition Mechanisms Elena Evguenieva-Hackenberg / (Peter Libby / Moshe Arditi / (Valerian Kagan / Eric Baehrecke) Charles Daniels / Mohammed Ouhammouch / Jeanine D’Armiento / Peter Libby / • Phagocyte Engulfment Mechanisms Felicitas Pfeiffer) Mark B. Pepys / Rama Natarajan / (Michael Hengartner / Kodi Ravichandran / • DNA Replication, Repair & Recombination Dan Simon) Simon Brown) (Malcolm White / Stephen Bell / • Thrombotic and Hemostatic Mechanisms • Alternative Phagocytes Takehiko Nohmi / Francesca Pisani / (Shaun R. Coughlin / Shaun R. Coughlin / (Kodi Ravachandran / Raymond Birge / Jeffrey Thorston Allers) Mark B. Taubman / Bruce Furie) Curtis) • Apoptotic Cell Recognition and Disease (Giovanna Chimini / Silvia Finnemann / Marcello Barcinski)

visit the frontiers of science: www.grc.org • Lipoprotein Metabolism • Condensed Matter Connections BIOLOGICAL MOLECULES IN THE GAS PHASE (Mary G. Sorci-Thomas / Miranda van Eck / (Anders Sorensen / Rob Shoelkopf) BATES COLLEGE Daniel J. Rader / Gwendalyn Randolph / • Precision Measurements LEWISTON, ME Alan M. Fogelman / Michael J. Thomas / (Larry Hunter / Dmitri Budker / Eric Hessels) JUL 24-29, 2005 Vasanthy Narayanaswami) • Ultrafast and High Field Physics MATTANJAH DEVRIES, CHAIR • Metabolic Syndrome and Atherosclerosis (Todd Ditmire / Margaret Murnane) ALBERT HECK, VICE CHAIR (Doug Vaughn / Peter Tontonoz / • Quantum Control Tony Ferrante) (David Tannor / Robert Jones / • Protein Misfolding and Formation • Macrophage Biology Dieter Meschede / Paul Haljan) (Mike Bowers / David Teplov / (Martha K. Cathcart / Catherine C. Hedrick / • Applications Alison Ashcroft / Andrea Sinz) Elaine Raines / William Muller / Ara Aslanian) (David Pritchard / Naomi Halas) • Folding and Conformation • Vascular Wall Stem Cell Biology (David Pratt / Joan-Emma Shea / (Elizabeth G. Nabel / Victor Dzau / ______Sven Hovmoller / Martin Jarrold) Shanin Rafii) • Single Molecules • Cell Biology of Atherosclerosis (Gilad Haran / X. Sunney Xie / (Joseph Loscalzo / Bradford C. Berk / Martin Zanni / Sander Woutersen) BARRIER FUNCTION OF MAMMALIAN SKIN Cecelia M. Giachelli / Thomas Quertermous / • Spectroscopy I MOUNT HOLYOKE COLLEGE Dennis Bruemmer) (Tim Zwier / Lavina Snoek / SOUTH HADLEY, MA • Human and Animal Michel Mons / Isabelle Compagnon) AUG 7-12, 2005 Genetics of Atherosclerosis • Spectroscopy II STEVEN HOATH & (Aldons J. Lusis / Paivi Pajukanta / (Karl Kleinnermans / Anne Zehnacker-Rentien GERALD KASTING, CO-CHAIRS Jonathan Smith / Alan Attie) / Wei Kong / Andrzej L. Sobolewski) ANTHONY RAWLINGS & • Spectroscopy of DNA MIKE ROBERTS, CO-VICE CHAIRS ______(Jerry Spooner / Brian Kohler / Glake Hill / Seong Keun Kim) • Comparative Biology and Physics of the • Top Down Approaches to Epidermal Barrier Protein Structure I ATMOSPHERIC CHEMISTRY (Walt Holleran / Gopi Menon / Bob Lochhead) (Evan Williams / Christoffer Borchers / BIG SKY RESORT • Membrane Structures and Transporters Scott McLuckey / Kathryn Breuker) BIG SKY, MT (Peter Elias / Gerd Schmitz / Andreas Stahl / • Top Down Approaches to SEP 4-9, 2005 Lily Bourgignon / Johanna Brand) Protein Structure II DAVID FAHEY, CHAIR • Nanoscale Characterization of Skin (Albert Heck / Neil Kelleher / DOUGLAS WORSNOP, VICE CHAIR Permeability Ron Heeren / Don Hunt) (Mike Roberts / Johannes Nitsche / • Feature Lecture • Key Perspectives Daniel Blankschtein / Yuri Anissimov) (Fred McLafferty) (Kenneth Demerjian / / • Water and the Stratum Corneum Spyros Pandis) (Joachim Fluhr / Tony Rawlings / Peter Elias) • Measuring and Modeling Aerosols and • Novel Analytical Techniques ______Their Impact on Climate (Richard Guy / Samir Mitragotri / (Kimberly Prather / Jose-Luis Jimenez / Stephen Hendrix) Barbara Turpin / Yinon Rudich / • Imaging the Stratum Corneum BIOMATERIALS: Joyce Penner) (Phil Wertz / Lars Norlen / Marek Haftek / Stig BIOCOMPATIBILITY / TISSUE ENGINEERING • Aerosols and Clouds Ollmar / Roger Wepf) HOLDERNESS SCHOOL (Graham Feingold / John Seinfeld / • Hot Topics PLYMOUTH, NH Sonia Kreidenweis / Thomas Peter) (Manige Fartasch) JUL 31-AUG 5, 2005 • Urban Pollution, Ozone Formation, and • Follicular Penetration KEVIN HEALY, CHAIR Regional Air Quality (Juergen Lademann / Nina Otberg / Joke ANDRES GARCIA, VICE CHAIR ( / Fred Fehsenfeld / Min Shao / Bouwstra / Michael Bonner) Yutaka Kondo / Paul Crutzen / • Debate: Does TEWL Correlate to • Lessons Learned: Robert Harley / Joost de Gouw) Epidermal Barrier Properties? Where Do We Go from Here? • Laboratory Studies of Atmospheric (Randy Wickett / Joachim Fluhr / Bob Chilcott) (Kevin E. Healy / David F. Williams / Processes Buddy D. Ratner) (Neil Donahue / Edward Lovejoy / ______• Cell-Material Interactions Michael Pilling) (Andrés Garcia / Christopher Chen / • The Biosphere Dennis Discher / Joyce Wong) (Mary Anne Carroll / Elizabeth Holland) • Host Response-Patient Variability BIOINFORMATICS: • Measurements from Space (Jonathan Black / Steve Goldring / FROM PREDICTIVE MODELS TO INFERENCE (Anne Douglass / Daniel Jacob / Joshua J. Jacobs) COLBY COLLEGE Randall Martin) • In Vivo Function of Tissue Equivalents WATERVILLE, ME • Global Modeling (James M. Anderson / Tony Atala / AUG 7-12, 2005 (Jose Rodriguez / Mark Lawrence / Laura E. Niklason / Stephen F. Badylak) DREW ENDY, CHAIR Michael Trainer / Michael Prather) • Protein Adsorption EDWARD MARCOTTE, VICE CHAIR (Igal Szleifer / Erwin A. Vogler / Tom Horbett) • Materials-Dependent In Situ Tissue ______• What is Biological Information? Regeneration • Timescales of Biological Information (William Wagner / Jeffery A. Hubbell / Processing Kristi Anseth / David Mooney) ATOMIC PHYSICS • Environmental Information • Wound Healing in Context TILTON SCHOOL • Genetic Information (Elazer R. Edelman / Julia Babensee) TILTON, NH • Epigenetic Information • Advances in Biomaterials Science JUN 26-JUL 1, 2005 (D.W. Grainger / David Tirrell / ERIC CORNELL, CHAIR ______Mark W. Grinstaff / Heather Maynard) CHRIS MONROE, VICE CHAIR • Stem Cell Therapy - Hype or Reality • Quantum Information (Michael Chapman / Luming Duan / ______Alex Kuzmich / Dietrich Leibfried) • Bose-Einstein Condensation ( / Lene Hau / Massimo Inguscio / David Weiss) • Degenerate Fermi Gases (Wolfgang Ketterle / Christoph Salomon / Cindy Regal)

visit the frontiers of science: www.grc.org BIOORGANIC CHEMISTRY CAG TRIPLET REPEAT DISORDERS CANCER MODELS & MECHANISMS PROCTOR ACADEMY MOUNT HOLYOKE COLLEGE BRYANT UNIVERSITY ANDOVER, NH SOUTH HADLEY, MA SMITHFIELD, RI JUN 12-17, 2005 JUL 24-29, 2005 JUL 24-29, 2005 KURT DESHAYES & NEIL MARSH, CO-CHAIRS MICHAEL LEVINE, CHAIR PIER PAOLO PANDOLFI, CHAIR LESLIE SLOAN & DIANE MERRY, VICE CHAIR RENE BERNARDS, VICE CHAIR PETER TONGE, CO-VICE CHAIRS • Clinical Presentation and Neuropathology • mRNA Translation Control and Cancer • Tinkering with the Central Paradigm (Sarah J. Tabrizi / Richard L.M. Faull / (David M. Sabatini / George Thomas / (Scott Silverman / Christine Chow / / Leslie Thompson) Lewis Cantley / Eric Holland) Jack Szostak) • Polyglutamine Impact on the Cytoskeleton • Nucleolar Network / Ribosome and Cancer • Nature Mimicked and Axonal Transport (Jacqueline Lees / Jason Weber) (Virgil Persec / Annelise Barron / (Larry Goldstein / Cynthia McMurray / • Aging, Telomeres and Cancer Alan Kennan / Stefan Matile) Kurt Fishbeck / Nancy Bonini) (Jerry Shay / ) • Nature Redesigned • Data Blitz and New Developments • Microenvironment and Metastasis (Trevor Douglas / Lynne Regan / (TBA - from posters / Jang-Ho Cha) (Harold Moses / Joan Massagué / Barrie Wilkinson) • Normal Function of Polyglutamine Proteins William Hahn) • Signals and Pathways and Impact on Pathogenesis • Mitotic Cell Cycle Control and Genomic (Alice Ting / Maurizio Pellecchia / (Randall N. Pittman / Sylvia Krobitsch / Instability Thomas Engler / Linda Hsieh-Wilson) Harry Orr / Al La Spada) (Prasad Jallepalli / Ashok Venkitaraman / • Enzymes and Biosynthesis • Neuronal Interactions in Polyglutamine Frederick Alt) (Mark Distefano / / Diseases and Excitotoxicity • Stem Cell and Cancer Stem Cell Craig Townsend) (Lynn Raymond / Paolo Calabresi / (George Daley / Michael Clarke / • Chemical Tools William Yang / Janet Dubinsky / Tetsuo Noda / Len Zon) (Tim Dore / David Berkowitz / Kerry Murphy) • Cooperative Tumorigenesis Bradley Smith / M.G. Finn) • Triplet Repeats in Non-Coding Regions / (Ron dePinho / Peter Sicinski) • Diseases and Therapies I RNA Mechanisms of Triplet Repeat Toxicity • Oncogenes, Tumor Suppressors and (Peter Toogood / Ranabir Sinha Roy / (Maurice S. Swanson / Peng Jin / Modifiers Christian Wiesman / Clifton Barry) Laura Ranum / Cheryl Wellington) (William Kaelin / René Bernards / • Diseases and Therapies II • Silencing RNAs Terry Van Dyke) (Celia Schiffer / Greg Weiss / Adrian Whittey) (Greg Hannon / Beverly Davidson / • Transcription, Epigenetic Control and • New Paradigms, New Technologies Philip Zamore / Neil Aronin / Nancy Wexler) Cancer (Henrik Pederson / Chad Mirkin / • Polyglutamine Species, Turnover and (Maarten Van Lohuizen / Nicholas La ) Autophagy Thangue) (David Rubinsztein / Steven Finkbeiner / ______Paul Patterson / Marcy MacDonald) ______• Preclinical, Clinical Trials and Development of Biomarkers (Rajiv Ratan / Bernhard Landwehrmeyer / BONES & TEETH CANNABINOID FUNCTION IN THE CNS Hindrik Mulder / Marie-Françoise Chesselet / KIMBALL UNION ACADEMY BATES COLLEGE Jenny Morton) MERIDEN, NH LEWISTON, ME • Keynote Plenary Address JUL 10-15, 2005 JUL 17-22, 2005 (Anne B. Young) RENE ST-ARNAUD, CHAIR KEN MACKIE & NEPHI STELLA, CO-CHAIRS PAMELA ROBEY, VICE CHAIR ______• Plenary Talk • New Developments (Gerard Le Fur) (Gérard Karsenty / Ronen Schweitzer / • Endocannabinoid Synthesis and Inactiva- Paolo Bianco) CALCIUM SIGNALLING tions • Transcriptional Control of Bone Cell THE QUEEN’S COLLEGE (Tung Fong / Daniele Piomelli / Ben Cravatt / Differentiation and Function OXFORD, UK Alex Makriyannis) (Patricia Ducy / Ernestina Schipani / JUL 24-29, 2005 • Cannabinoid Receptors and Signal Hiroshi Takayanagi / Donald Glass) COLIN TAYLOR, CHAIR Transduction • Advances in Stem Cell Biology: INDU AMBUDKAR, VICE CHAIR (Manuel Guzman / Deborah Lewis / From the Inner Cell Mass to the Tamas Freund / Jean Antoine Girault) Post-Natal Organism • Spatial Organisation of Ca2+ Signalling • Cannabinoids and Neuronal Plasticity I (Pamela G. Robey) (Alexei Tepikin / Mary Kennedy / (Wade Regehr / Masanobu Kano / • Mineral Homeostasis and Calcification Gyorgy Hajnoczky / Pablo Castillo / Brad Alger) (L. Darryl Quarles / Robert Terkeltaub / Clara Franzini-Armstrong) • Cannabinoids and Neuronal Plasticity II Lynda Bonewald / Paul Price / Dwight Towler) • Intracellular Ca2+ Channels (Jane Sullivan / Dan Feldman / • Teeth / Craniofacial Biology (Barbara Ehrlich / Noriaki Ikemoto / Sacha Nelson / David Lovinger) (Marc McKee / Irma Thesleff / Paula da Fonseca / ) • Adaptive Changes with Chronic Cannab- Michael Paine / Songtao Shi) • Ca2+ Regulation of Channels inoid Use • Signaling / Cytokines (Susan Hamilton / Gerda Breitwieser / (Phil Iredale / Laura Sim-Selley / (Matthew Gillespie / Robyn Starr / David Yue / Kevin Foskett) Olvier Manzoni / Stan Thayer) Toshiyuki Takai / Roland Baron) • Ca2+ Entry • Food, Aversion, Reward, Sex, and Pain I • Osteoclast / Bone Resorption (Jim Putney / Michael Tymianski / (Carl Lupica / George Kunos / Beat Lutz / (Robert L. Jilka / Brendan Boyce / Victoria Bolotina / ) Rafael Maldonado) Stavroula Kousteni / Mitch Schaffler) • TRP Proteins • Food, Aversion, Reward, Sex, and Pain II • Skeletal Anabolic Agents (Bernd Nilius / Michael Caterina / (George Nomikos / Don Simone / (Nicola C. Partridge / Sundeep Koshia / Veit Flockerzi / Richard Sandford) Aron Lichtman) Lin Qin / Ross Garrett / Peter Bodine) • Ca2+ and Secretion • Interactions with Other Signaling Systems • Keynote Lecture (Indu Ambudkar / Ege Kavalali / (Andrea Giuffrida / Virginia Pickel / (René St-Arnaud / Henry Lee) Pete Thorn / Shmuel Muallem) Lakshmi Devi) • Ca2+ and Vascular Function ______(Mike Berridge / Mark Nelson / ______Lother Blatter / Dirk Van Helden) • Decoding Ca2+ Signals (Manuela Zaccolo / Ian Parker / Bob Burgoyne / ) • Plenary Lecture (Katsuhiko Mikoshiba)

visit the frontiers of science: www.grc.org CARBOHYDRATES CATECHOLAMINES • Global Analyses of Metals in Cells TILTON SCHOOL PROCTOR ACADEMY (David Eide / Tom O’Halloran / TILTON, NH ANDOVER, NH Simon Andrews / Mary Lou Guerinot) JUN 19-24, 2005 JUL 24-29, 2005 • Metallochaperones and Intracellular GEERT-JAN BOONS, CHAIR DAVID SULZER, CHAIR Trafficking TODD LOWARY & JILL BECKER, VICE CHAIR (Tom O’Halloran / Val Culotta / PENG WANG, CO-VICE CHAIRS Nigel Robinson / Jonathan Gitlin) • Behavior and Neurotransmission • New Synthetic Methods for the Synthesis (Stephanie Cragg / ______of Complex Oligosaccharides Keynote Lecture: ) (Geert-Jan Boons / Alexei Demchenko / • Driving Neuronal Activity Takashi Takahashi / Andrea Vasella) (R. Mark Wightman / Suzanne Haber / CELL CONTACT & ADHESION • New Directions in the Target Synthesis of Susan Sesack / Peter Redgrave / PROCTOR ACADEMY Complex Oligosaccharides and J. Paul Bolam) ANDOVER, NH Glycoconjugates • Is Dopamine the Substrate for Reward? A JUN 26-JUL 1, 2005 (George Peng Wang / Debate GERALD GRUNWALD, CHAIR Jacqueline Gervay-Hague / Zhongwu Guo / (Jill Becker / Nora Volkow / ELISABETTA DEJANA, VICE CHAIR Gijs van der Marel) P. Read Montague / Kent Berridge / • Design, Synthesis and Biological John Salamone) • Cadherins in Perspective Evaluation of Inhibitors of Carbohydrate • Synaptic Modulation (Masatoshi Takeichi / Rolf Kemler) Processing Enzymes (D. James Surmeier / Patricio O’Donnell / • Cadherin Genomics and Structural Biology (Todd Lowary / B. Mario Pinto / John Williams / Jose Bargas) (Deborah Leckband / Rheinhard Gessner / Vern Schramm) • Neurodegenerative Mechanisms Mitsuhiko Ikura) • Genomics, Proteomics and the New Era of (Serge Przedborski / William Dauer / • Cadherins and Cell Signaling Glycomics Ana Maria Cuervo / ) (Christopher Chen / Stephen Byers / (Ole Hindsgaul / Catherine Costello / • Receptors / Second Messengers Barry Gumbiner) James Paulson / Nicola Pohl) (Robert Edwards / Nigel Bamford / • Cadherins and Receptor Interaction • Carbohydrate Bio-Engineering Ivan Diamond / Jeremy Seamans) (Rachel Hazan / Robert Brackenbury / (Antoni Planas / John Thorson / • Workshop: Non-Neurotransmitter Nikolaos Robakis) Stephen Withers) Catechols • Cadherin-Integrin Crosstalk • Stuctural Studies of Oligosaccharides and (Okezie Aruoma / Luigi Zecca) (Jack Lilien / Keith Johnson / Carbohydrate Binding Proteins • Behavioral Disorders Maria Kukuruzinksa) (Monica Palcic / Daan van Aalten) (Michael Zigmond / Terry Robinson / • Cadherins and Development I • Complex Carbohydrates and Microbes Antonello Bonci / Richard Palmiter) (Walter Birchmeier / James Nelson / Elisabetta (David Bundle / Hung-Wen Liu / • Transporters and Presynaptic Regulation Dejana) Stefan Oscarson / Suzanne Walker) (Nancy Zahniser / Jonathan Javitch / • Cadherins and Cancer • Chemistry and Biochemistry of Glycopro- Aurelio Galli / Mark Caron / (Andrea McClatchey / Al Reynolds / teins and Glycopeptides Annette Fleckenstein / Mark von Zastrow) Pamela Cowin) (Horst Kunz / Yukishige Ito) • Workshop: Discussion Group • Cadherins and Development II • Glyconanotechnology (Michael Zigmond) (Christoph Redies / (Nicolai Bovin / J.P. Kamerling / • Kopin Award Lectures Juliet Daniel / Jeffrey Hardin) J. ) (to be selected from junior investigators) • Hot Topics • Recognition of Carbohydrates by Artificial • Integrating Synapses with Behavior Receptors (Anne Etgen / Regina Carelli / (Amit Basu / Anthony Davis / Binghe Wang) Plenary Lecture: Paul Greengard) ______CELL PROLIFERATION, MOLECULAR & GENETIC BASIS OF COLBY-SAWYER COLLEGE CATCHMENT SCIENCE: INTERACTIONS OF CELL BIOLOGY OF METALS NEW LONDON, NH HYDROLOGY, BIOLOGY & GEOCHEMISTRY BATES COLLEGE JUN 26-JUL 1, 2005 COLBY COLLEGE LEWISTON, ME MICHAEL TYERS, CHAIR WATERVILLE, ME JUL 3-8, 2005 MICHAEL YAFFE, VICE CHAIR JUL 17-22, 2005 N ROBINSON & DENNIS WINGE, CO-CHAIRS CHRISTINE ALEWELL & ANDREW DANCIS, VICE CHAIR • Keynotes DOUGLAS BURNS, CO-CHAIRS (Gerard Evan / Steve Elledge) ELIZABETH BOYER, VICE CHAIR • Metal Homeostasis in the Central Nervous • Growth System (Bruce Edgar / Mike Hall / Morrie Birnbaum / • Ecosystem Disturbance (Jonathan Gitlin / John Burn / Leah Harris / George Thomas / Laura Johnston) (Peter Groffman / Dale Johnson / Tracey Rouault) • Signaling Lindsay Rustad / Ralph Boerner / • Transport of Metal-Complexes ( / Dafna Bar-Sagi / Ben Neel / Beate Michalzik / Klement Tockner / (Jerry Kaplan / Jon Barasch / Helen McNeill) Keith Eshleman) Carolyn Philpott / Andy McKie / Mark Fleming) • Cell Cycle • Ecosystem Restoration • Membrane Metal-Transport (Orna Cohen-Fix / Wade Harper / (David Allan / Louise Heathwaite / (Nancy Andrews / Dennis Thiele / Helena Richardson / Matthias Peter / Emily Bernhardt / John Quinn / Bob Harris / Svetlana Lutsenko / Dave Eide) Joan Ruderman) Joy Zedler / Bob Foy) • Metal-Responsive Transcription and • Chromosome Replication and Dynamics • Field Trip: Acadia National Park Signaling ( / Vicki Lundblad / Catchments (David Giedroc / Amanda Bird / Simon Labbe / John Diffley / Dan Durocher) (Steve Kahl) Simon Whitehall / Walter Schaffner) • Checkpoints • Long-Term Monitoring and Experimental • Post-Transcriptional Metal-Responses (Helen Piwnica-Worms / Sally Kornbluth / Manipulation (Betty Leibold / William Walden / Carol Prives / Susan Lees-Miller / Mike Yaffe) (Heleen de Wit / Colin Neal / Mick Petris / Glen Andrews) • Systematics and Mathematical Modeling Christine Goodale / Richard Skeffington / • Metals in Organelles (Fred Cross / Ravi Iyengar / Jill Baron / Peter Dillon / Michel Meybeck) (Val Culotta / Andy Dancis / Rebecca Heald / Mike Snyder) Jerry Kaplan / Roland Lill / John Weiss) • Cancer • Intercellular Metal-Homeostasis ______(Tak Mak / Jackie Lees / Jean Wang) (Jim Kushner / Sophie Vaulont / • Gene Regulation Nancy Andrews / Tom Ganz) (Dan Gottschling / Nick Dyson / Bill Tansey / Ali Shilatifard)

visit the frontiers of science: www.grc.org CELLULAR OSMOREGULATION: SENSORS, • Ferroelectrics: • Interface Characterization TRANSDUCERS AND REGULATORS Micromechanics and Fatigue (Richard Colton / Andreas Engel / SALVE REGINA UNIVERSITY (Michael Hoffmann / Jurgen Rödel / Rachel McKendry) NEWPORT, RI Norman Fleck) • Optical Chemical Sensor Technology AUG 7-12, 2005 • Nanoindentation of Semiconductor and (Reinhard Niessner / Jerome Faist / D. WAYNE BOLEN & IAN BOOTH, CO-CHAIRS Ceramic Materials Jiri Homola / Karl Unterrainer) RAINER HEDRICH, VICE CHAIR (Bill Clegg / Jody Bradby / Robert Cook) • News on Electrochemical Sensors • Nanocomposites and Nanomechanical (Ingemar Lundström / Dave Williams / • Cellular Osmoregulation Properties of Ceramics Mark Lonergan) (Maurice Burg / Janet Wood) (Raj Bordia / Giuseppi Pezzotti / Bill Curtin) • in Chemical Sensing • Stress Response: Sensing and Response • Fabrication of MEMS Structures (Andreas Hierlemann / Phaedon Avouris / Integration (Rob Dorey / Mark Spearing) James Heath / Reginald Penner) (Karlheinz Altendorf / Kazuo Shinozaki / • Novel Fabrication and Synthesis of • Short Poster Talks Dietmar Kültz) Inorganic Materials (Anthony Coleman) • Cell Volume Regulation (John Halloran / Jennifer Lewis / • New Frontiers (Donald Hilgemann / Paul Blount / Rajeesh Naik) (Elizabeth Hall / Sylvia Daunert / Samantha Miller) • Structure/Property Relations in Biological Kevin Linker / Jeffrey Borenstein) • Macromolecule-Water-Solute Interactions Materials (Valarie Daggett / Gary Pielak / (Bob McMeeking / Adrian Mann / ______Daniel Harries / Jörg Rösgen) Robert Ritchie) • Osmotic Stress: Organism Responses • Materials in Biomedical Applications (Richard Morimoto / Volker Müller / Jay Gralla) (Brian Lawn / Mike Swain) CHEMISTRY EDUCATION • Signal Transduction and Osmoregulation • Thursday Night After Dinner Speaker RESEARCH & PRACTICE (Masayori Inouye / Haruo Saito / (Brian Derby / TBA) CONNECTICUT COLLEGE Julian Schroeder) NEW LONDON, CT • Aquaporins ______JUN 26-JUL 1, 2005 (Mark Knepper / Christophe Maurel / STACEY LOWERY BRETZ, CHAIR Andreas Engel) CHRISTOPHER BAUER, VICE CHAIR • Osmolyte Accumulation and its Impact on CHEMICAL OCEANOGRAPHY Membrane Structure & Stability TILTON SCHOOL • Shaping the Research Agenda in Chemistry (Joseph Zacchai / Antoinette Killian / TILTON, NH Education Dennis Dougherty / Fred Sachs) AUG 7-12, 2005 (Art Ellis / / George Bodner) • Integrated Function and Osmoregulation CLARE REIMERS, CHAIR • Mentors & Teachers: The Transition States (Erhard Bremer / Kevin Strange) EDWARD BOYLE, VICE CHAIR of Our Profession (Mary Atwater / Kate Scantlebury / ______• CO2 Induced Changes in Ocean Calcifica- Isiah Warner / Donald Wink) tion • Learning in the Chemistry Laboratory (Ken Caldeira / Victoria Fabry / (Norbert Pienta / Dawn Rickey / CELLULASES & CELLULOSOMES Joanie Kleypas) Brian Woodfield) PROCTOR ACADEMY • Land-Ocean Interactions • Advances in Measuring Student Learning ANDOVER, NH (Sybil Seitzinger / Wei-Jun Cai / (Thomas Holme / Melanie Cooper / AUG 7-12, 2005 James Bauer / Thomas Bianchi) Diane Ebert-May / Angelica Stacy) TUULA TEERI, CHAIR • Science from Sensors • Learning On-Line: Facilitating Student R. ANTONY WARREN, VICE CHAIR (Ronnie Glud / Marylou Tercier-Waeber / Understanding Stefan Hulth) (Jimmy Reeves / Marcy Hamby Towns / • Keynote Lectures • Coastal -Open Ocean Connections Gregor Novak) (Rajaj Atalla / Al Boraston) (Chen-Tung Arthur Chen / Denis Gilbert / • Visualization & Language in Learning • Cellulose and Cellulose-Binding Modules Burke Hales / Nicolas Gruber) Chemistry (Harry Gilbert / Paul Knox) • Oceanic Iron: Concentrations and Isotope (Marcia Linn / Guy Ashkenazi / Roy Tasker / • Enzyme Producers: Physiology and Compositions Maria Oliver-Hoyo) Genomics (Robert Sherrell / Kenneth Johnson / • The Chemistry of History & The History of (Joel Cherry / Merja Penttilä) Silke Severmann) Chemistry • Enzyme Mechanisms and Engineering • Anaerobic Biogeochemistry (Zafra Lerman / Pierre Laszlo / (Marc Cleayssens / Anu Koivula / Bill Adney) (Mary Scranton / Marcel Kuypers / George Mary Virginia Orna) • Plant Cellulases Luther / Josef Werne) • Student-Centered Learning: The Role of (David Wilson / Emma Master / • Crustal Recycling and Fluxes Faculty Development Breeanne Urbanowicz) (Michael Mottl / Terry Plank / Geoff Wheat) (Amy Phelps / Jennifer Lewis / • Cellulosomes: Structure, Function and • Biomolecules, Speciation and Maureen Scharberg) Engineering Paleo-Proxies • Disseminating Pedagogical Change: (Ed Bayer / Roy Doi) (Roger François / James Moffett / Peter Increasing Our Percent Yield • Enzymatic Processing of Cellulose Santchi / Gideon Henderson) (Chris Bauer / Rick Moog / (Mike Himmel / John Tomashek) • Reconstructing the Chemistry of Past Pratibha Varma-Nelson) • Other Polysaccharidases Oceans (Colin Mitchinson / Leila LoLeggio / (Ed Boyle / Rebecca Robinson / Julian Sachs) ______Maria Hrmova) ______CHROMOSOME DYNAMICS COLBY-SAWYER COLLEGE CHEMICAL SENSORS & INTERFACIAL DESIGN NEW LONDON, NH CERAMICS, SOLID STATE STUDIES IN THE QUEEN’S COLLEGE JUL 31-AUG 5, 2005 TILTON SCHOOL OXFORD, UK ABBY DERNBURG, CHAIR TILTON, NH AUG 28-SEP 2, 2005 N. PATRICK HIGGINS, VICE CHAIR JUL 17-22, 2005 BORIS MIZAIKOFF, CHAIR BRIAN DERBY, CHAIR ANTHONY COLEMAN, VICE CHAIR • Recombination and Genome Plasticity JOHN BLENDELL, VICE CHAIR (David Sherratt / Nancy Kleckner / • Molecular Recognition John Moran / Susan Lovett) • Failure Mechanisms of Si and Devices (Boris Mizaikoff / Jean-Marie Lehn / • Links Between Chromosome Structure and (Roberto Ballarini / Zhigang Suo) Christoph Fahrni) Function • Interface Architectures (John Marko / John Battista / (Tony Ricco / Mark Grinstaff / Nancy Maizels / Nick Cozzarelli / Jerry Atwood / Günther Tovar) Ling Juan Wu)

visit the frontiers of science: www.grc.org • Evolution of Genomes and Chromosomes CLUSTERS, NANOCRYSTALS & COASTAL OCEAN CIRCULATION (Matt Meselson / L. Aravind / Evan Eichler / NANOSTRUCTURES COLBY-SAWYER COLLEGE Fernando Pardo Manuel de Villena) CONNECTICUT COLLEGE NEW LONDON, NH • Chromosome Sites with Special Functions NEW LONDON, CT JUN 5-10, 2005 (Julie Cooper / Arshad Desai / Don Cleveland / JUL 31-AUG 5, 2005 STEVEN LENTZ, CHAIR Hiro Funabiki / Kelly Dawe / Gary Karpen / MOUNGI BAWENDI & JAMES O’DONNELL, VICE CHAIR George Chaconas) ORI CHESHNOVSKY, CO-CHAIRS • Cell Division and Chromosome Segrega- A. WELFORD CASTLEMAN, JR. & • Dispersal of Terrestrial Runoff tion VICKI COLVIN, CO-VICE CHAIRS (James O’Donnell / John H. Simpson / (Barb Funnell / Sue Biggins / Kenn Gerdes / Robert Chant) Stuart Austin / Jason Swedlow / • Opening Session • Shelf Circulation and Shelf Edge Pro- Daniela Cimini / Rebecca Heald) (Peidong Yang / Daniel Neumark) cesses • Meiosis and Sporulation • Electronic Properties (Robert Beardsley / John S. Allen / (Scott Hawley / Yasushi Hiraoka / ( / Bernd von Issendorff / Glen Gawarkiewicz / Edward Dever) Sigal Ben-Yehuda) Uri Banin / Atsushi Nakajimi) • High Latitude Shelf Processes • Organization and Dynamics of Transcrip- • Magnetism (Tom Weingartner / Robert Pickart / tional Domains (Lai-Sheng Wang / Walt de Heer) John Klinck) (Barbara Meyer / Barbara Panning / • Optical Properties • Physical Influences on Dmitry Vassylyev / Shiv Grewal) (Alex Zunger / Victor Klimov) Coastal Ecosystems • Condensation and Cohesion • Structure (Richard Signell / Jonathan Sharples / (Tatsuya Hirano / Rolf Jessberger / (Younan Xia / Martin Jarrold) Andy Visser / David Townsend) ) • Nanostructures and Biology • Nearshore Processes • Replication and the Cell Cycle (Amit Meller / Christine Keating / (Gail Kineke / Peter Traykovski / (Ken Kreuzer / Alan Grossman / Itamar Wilner) Tuba Ozkan-Haller) Steve Bell / Susan Forsburg) • Physical Properties • Estuarine and (Very) Shallow Water (Hellmut Haberland / Uzi Landman) Dynamics ______• Functional Structures (Parker MacCready / James Lerczak / (Hongjie Dai / Eran Rabani / James Heath) Mark Stacey / Heidi Nepf) • Photonics • Coastal Interactions with the Atmosphere (Hongkun Park / Naomi Halas / Yoel Fink) (Roger Samelson / James Edson / John Bane) CHRONOBIOLOGY • Vertical Mixing SALVE REGINA UNIVERSITY (Tom Rippeth / Jonathan Nash / NEWPORT, RI ______Jennifer MacKinnon / Eric D’Asaro) JUL 31-AUG 5, 2005 • Scientific Insights from MARTHA GILLETTE, CHAIR Coastal Observatories TILL ROENNEBERG, VICE CHAIR CO ASSIMILATION IN PLANTS: 2 (Steven Lentz / Jack Barth / Clinton Winant) GENOME TO BIOME • The Whole Cell Clock CENTRE PAUL LANGEVIN (Gene Block / Martha Merrow / AUSSOIS, FRANCE ______Gerald Pollack / Howard Petty / SEP 11-16, 2005 Masayuki Ikeda) GEORGE BOWES & • Oscillator Networks: Gene and Cell SUSANNE VON CAEMMERER, CO-CHAIRS COATINGS & FILMS Systems ANNE BORLAND & COLBY-SAWYER COLLEGE (Carla Green / Paolo Sassone-Corsi / MICHAEL SALVUCCI, CO-VICE CHAIRS NEW LONDON, NH Deb Bell-Pedersen / Satchin Panda / JUL 10-15, 2005 Shelley Tischkau / Hitoshi Okamura) • Conference Opening DOUGLAS WICKS, CHAIR • Oscillator Networks: Cells and Tissues (George Bowes / Susanne von Caemmerer) WILLIAM SIMONSICK & • (Robert Y. Moore / Laura Smale / Erik Herzog Photosynthesis in a Higher CO2 World JUDITH STEIN, CO-VICE CHAIRS / Horacio de la Iglesia / Rae Silver) (Howard Griffiths / Gregory Asner / • Entrainment: Molecular Sensors to Ulf Riebesell) No information available at press time. Please • Behavioral Change CO2 Entry check the GRC web site for details. (Russell Foster / Susan Golden / (James Moroney / Stephen Maberly / Charalambos Kyriacou / Benjamin Tu / Fiona Woodger / Yuko Hanba) ______Steven Kay / / • Stomatal Regulation and Metabolism Till Roenneberg) (Keith Mott / Sarah Assmann / William Outlaw) • Clocks, Sleep and Genes • Regulation of CO -Assimilating Enzymes 2 COLLAGEN (Joan Hendricks / Ketema Paul / Fred Turek / (Martin Parry / Inger Andersson / COLBY-SAWYER COLLEGE Giulio Tonini / Paul Shaw) Archie Portis / Jean Vidal) NEW LONDON, NH • Clocks in Translation • Chloroplastic Carbon Metabolism JUL 24-29, 2005 (Ann-Marie Chang / Phyllis Zee / (Christine Raines / Brigitte Gontero-Meunier / PETER BRUCKNER, CHAIR Liz Maywood / Joseph Bass / Marina Antoch / Alison Smith) • DAVID BIRK, VICE CHAIR Cheng Chi Lee / Scott Davis) C4 and CAM Photosynthesis • Output Signals and Peripheral Clocks (Anne Borland / Julian Hibberd / • New Collagens (Elizabeth Klerman / Paul Taghert / Gerald Edwards / John Cushman) (Florence Ruggiero) Amita Sehgal / Ruud Buijs / Charles Weitz) • Early-Career Scientists and “Hot-Off-The- • Expression, Biosynthesis and Regulation • Temporal Aspects of Press” Short Talks of Collagens Behavioral Integration (Michael Salvucci) (Johanna Myllyharju) (Carolina Escobar / David Weaver / • Emerging Technologies • Protease Processing of Matrix Michaela Hau / Joel Elmquist / (Hans Bohnert / Itzhak Kurek / Sacha (Suneel Apte) Nicholas Mrosovsky / Donald Pfaff / Baginsky / Sylvie Lalonde / Alisdair Fernie) • Supramolecular Matrix Assemblies Martin Ralph) • After-Banquet Featured Speaker (Paul Bornstein) • Visions of Synthesis (Susanne von Caemmerer / Mark Stitt) • Cell Adhesion to Matrix - Cryptic Adhesion (Mick Hastings / David Welsh / (Johannes Eble) Patricia Lakin-Thomas / Joseph Takahashi / ______• Collagen Diseases ) (Leena Ala-Kokko) • Gene Therapy and Regenerative Medicine ______(Leena Bruckner-Tuderman) • Invertebrate and Vertebrate Animal Models (Kathy Cheah) • Hot Topics in the Field

visit the frontiers of science: www.grc.org COMBINATORIAL & HIGH THROUGHPUT • Chemical Genetics • Bulk Technologies - I MATERIALS SCIENCE (Kip Guy / Anna Mapp / Young-Tae Chang / • Emerging Technologies THE QUEEN’S COLLEGE Tom Kodadek) • Special Session: OXFORD, UK Near and Far-Range Standoff Detection AUG 14-19, 2005 ______• Bulk Technologies - II JAMES CAWSE & • Trace Technologies - II: Detection WILHELM MAIER, CO-CHAIRS • Challenges for the Future: DAVID ROTHMAN & An International Perspective COMPUTER AIDED DRUG DESIGN ULRICH SCHUBERT, CO-VICE CHAIRS TILTON SCHOOL TILTON, NH ______• How to Handle Diversity - Biomaterials and JUL 31-AUG 5, 2005 Enzymes PETER JURS, CHAIR (Oleg Kolosov / Manfred T. Reetz / RICHARD LEWIS, VICE CHAIR DEVELOPMENTAL BIOLOGY Bruce Eaton) PROCTOR ACADEMY • Theoretical Aspects of Materials Discovery • Scoring Docking Poses ANDOVER, NH (James Cawse / Dimitris Agrafiotis / (Michal Vieth / Chris Murray / JUN 19-24, 2005 Francois Gilardoni / Mikk Lippmaa) Arthur Doweyko) RICHARD HARLAND, CHAIR • Discovery of Knowledge • Cheminformatics I STEPHEN COHEN, VICE CHAIR (Claude Mirodatos / Ferdi Schueth / (Mik Lajiness / Gavin Harper / Fred Hamprecht) Val Gillet / Mark Mackey) • Asymmetry and Cell Migration • Materials Development • Cheminformatics II (Richard Harland / Chris Doe / Rueyling Lin / (Eric Amis / Y. Matsumoto / K.-S. Sohn / (Dan Ortwine / Bob Clark) Ruth Lehmann / Chris Wylie / ) Oden Warren) • Energetics of Molecular Recognition • Genomic and Genetic Approaches (I) • Search for Novel Materials (Andy Good / Ernersto Frieire / (Ruth Lehmann / Julie Ahringer / (Ichiro Takeuchi / Charles Olk / Seong Woo) Celia Schiffer / Adrian Elcock) Norbert Perrimon / Makoto Furutani-Seiki / • Industrial Experiences with HTE • Modelling GPCRS Ron Plasterk / ) (William Flanagan / Jennifer Holmgren / (Antonia do Amaral / Caterina Bissantz) • Evolution, Morphogens and Modeling Thomas Brinz / • Protein Modelling (Julie Ahringer / Mike Levine / Naama Barkai / Victor Adamian / Katharine Allen) (Brian Shoichet / Holger Gohlke / Andrej Sali) Robb Krumlauf / Nipam Patel) • Discovery and Optimization of • State of the Nation • Growth and Patterning Heterogeneous Catalysts (Tudor Oprea / Garland Marshall) (Janet Heasman / Laura Johnson / (Dirk Demuth / Pierre Jacobs / Peter Claus) • New Technologies Matt Freeman / Joel Rothman / • Discovery and Optimization of (Tom Fox / Jon Mason / Nicolas Froloff) David Wilkinson / Detlef Arendt) (Ulrich Schubert / Rolf Muelhaupt / • Case Studies • Cell Migration and Boundary Formation Mark Bradley / Kathryn Beers / (Bill Egan / Mike Sabio) (Elizabeth Robertson / Chris Stafford) Marianne Bronner-Fraser / Pernille Rørth / • Discovery and Applications of Olivier Pourquie / Ray Keller) Homogeneous Catalysts ______• Signaling and Pattern Formation (M.G. Finn / Olivier Lavastre / (Pernille Rørth / / Albrecht Berkessel) Christoph Niehrs / Phil Soriano / CONDENSED MATTER PHYSICS Arthur Lander / Gail Martin / Alex Schier) CONNECTICUT COLLEGE ______• Growth and Pattern Formation (II) NEW LONDON, CT (Marianne Bronner-Fraser / Liz Robertson / JUN 19-24, 2005 Lilianna Solnica-Krezel / Ernst Hafen / PAUL MCEUEN, CHAIR COMBINATORIAL CHEMISTRY Iswar Hariharan) ROBERT MAGERLE, VICE CHAIR PROCTOR ACADEMY • Signaling and Morphogenesis During ANDOVER, NH Organogenesis • Biology Meets Materials AUG 21-26, 2005 (Laura Johnson / Janet Heasman / (Joanna Aizenberg) SAMUEL GERRITZ, CHAIR Andy McMahon / Scott Fraser / • Colloids as Molecules, R. KIP GUY, VICE CHAIR Richard Harland / Doug Melton) Liquids, and Solids • Growth and Pattern (III) (David Weitz / David Pine) • Target Class Libraries (Stephen Cohen / Gary Ruvkun / • Membrane Interfaces and Networks (Christine Brotherton-Pleiss / Hartmuth Kolb / Cliff Tabin / Stephen Cohen) (Jay Groves / Owe Orwar) David Drewry / Jon Ellman) • Novel Manipulation and • Case Studies I Imaging Techniques ______(Daryl Sauer / Brian McKittrick / (Robert Westervelt / Lois Pollack) Rachael Hunter / Guy Breitenbucher / • Nano Meets Bio Craig Lindsley / Stefan Warner) (Serge Lemay / Luke Lee / Michael Roukes) DRUG METABOLISM • Library Synthesis Capacity: Rent or Buy? • Vortices HOLDERNESS SCHOOL (John Porco / James Connelly / (David Nelson / Cynthia Olson Reichardt) PLYMOUTH, NH Libing Yu / Bill Coates) • Synthesis JUL 10-15, 2005 • Libraries Based on Natural Products (Rustem Ismagilov / Tom Mallouk) LARRY WIENKERS, CHAIR (Sam Gerritz / Takashi Takahashi / • Future of Condensed Matter Physics LESLIE BENET, VICE CHAIR Gunda Georg / Ganesan / Annaliese Franz / (Sid Nagel) Scott Schaus) • Advances in the Basic Knowledge of • CMLD Overview Cytochrome P450 Towards the Understand- (John Schwab / John Porco / Stefan Werner / ______ing of Drug Metabolism: Part 1 - Structure/ Jeff Aube / Harvard) Function • Synthesis and Screening of Biopolymers (Stephen G. Sligar / Michael R. Waterman / (Shelli McAlpine / Peter Seeberger / DETECTING ILLICIT SUBSTANCES: Ilme Schlichting / Eric F. Johnson / Laura Kiessling / Alice Ting / Akira Kawamura) EXPLOSIVES & DRUGS Richard N. Armstrong) • Novel Solid- and Solution-Phase Synthetic LES DIABLERETS CONFERENCE CENTER • Advances in the Basic Knowledge of Methods LES DIABLERETS, SWITZERLAND Cytochrome P450 Towards the Understand- (John Quinn / Rolf Breinbauer / AUG 28-SEP 2, 2005 ing of Drug Metabolism: Part 2 - Oxidation Eric Jacobson / Mark Bradley) RICHARD LAREAU & JEANNE LIN, CO-CHAIRS Reactions • Case Studies II (William M. Atkins / Jeffrey P. Jones / (Richard Austin / Jeremy Green / • Policy Discussion: Sason Shaik / Stephen G. Sligar) Anna Pendri / Christopher Hulme / Privacy Versus Security Ivan Lorkovic) • Trace Detection Technologies - I: Sampling and Sensing & Detection • Olfactory and Detection R&D

visit the frontiers of science: www.grc.org • Drug Metabolism-Consequences to Toxicity • Elastic Tissue Development ELECTRONIC MATERIALS, CHEMISTRY OF / Pharmacology (Robert Mecham / Charles Little / CONNECTICUT COLLEGE (Judy Bolton / Jim Fishbein / Natalia Evan Zamir / Eiichi Hirano) NEW LONDON, CT Tretyakova / Gregory R.J. Thatcher / • Lysyl Oxidases JUL 17-22, 2005 John M. Essigmann) (Katalin Csiszar / Robert Rucker / CHERIE KAGAN & • Expression and Tissue Distribution of Drug Herb Kagan / Philip Trackman / THOMAS MALLOUK, CO-CHAIRS Metabolizing Enzymes Pascal Sommer / Tiansen Li / Ian Hornstra) CHRISTOPHER CHIDSEY & (Jeffery C. Stevens / J. Steven Leeder / Xinxin • Elastic Fiber Molecules - GARY TAYLOR, CO-VICE CHAIRS Ding / Ronald N. Hines) Knock-Out Models • Integrating Preclinical Information to (Elaine Davis / Michael Shipley / • Nanocrystal Materials and Devices Predict Human Drug Disposition Giorgio Bressan / Jouni Uitto / Luca Carta) (Cathy Murphy / Alfons von Blaarden / (Timothy S. Tracy / Leslie Z. Benet / • Heritable Elastic Fiber Diseases Yang Yang) R. Scott Obach / Mary Paine / (Zsolt Urban / Dianna Milewicz / • Molecular Junctions J. Brian Houston) Geert Mortier / Edwin Stone / András Váradi / (Stuart Lindsay / James Kushmerick / Theresa • Innovative Clinical Approaches in Ivonne Ronchetti) Mayer / Mark Ratner) Underwriting Disposition Factors in Early • Elastic Fibers and Growth Factors • Frontiers of Electronic and Magnetic Drug Development (Lynn Sakai / Dan Rifkin / Vesna Todorovic / Materials / Extraordinary Posters (Paul G. Pearson / Scott Patterson / Gerhardt Sengle / Katri Koli / Bart Loeys) (Harry Atwater) Richard Hargreaves) • Degenerative Elastic Fiber Disorders • Organic Devices • Novel Technology Approaches to Applied (Bill Parks / Steve Shapiro / (Paul Barbara / George Malliaras / to Drug Discovery and Development Drug Marlene Rabinovitch / Robert Thompson / / Janos Veres) Metabolism Steve Shapiro / Charles Boyd / • Nanoelectronics (Kenneth R. Korzekwa / Daniel E. Murnick / Richard Pierce) ( / Joerg Appenzeller / William M. Atkins / Catherine Booth-Genthe) • Elastic Fiber Engineering and Biomaterials Lars Samuelson / Georg Duesberg) (Tony Weiss / Ivan Vesely / Fred Keeley / Alex • Biomolecular and Supramolecular ______Seifalian / Richard Black) Electronics (Samuel Stupp / Amy Blum) ______• Semiconductor Materials, Growth and Devices DYNAMICS AT SURFACES (Judy Hoyt / Jeff Calvert / Veena Misra / PROCTOR ACADEMY Robert Hicks) ANDOVER, NH ELASTOMERS, NETWORKS & GELS • Polymeric Materials and Patterning AUG 14-19, 2005 COLBY-SAWYER COLLEGE (Christopher Ober / Charles Black / GILBERT NATHANSON, CHAIR NEW LONDON, NH Ralph Dammel) BRET JACKSON, VICE CHAIR JUL 17-22, 2005 • Electronics at Interfaces GREGORY MCKENNA, CHAIR (Ellen Williams / Eric Garfunkel) • State-to-State Dynamics H. HENNING WINTER, VICE CHAIR (Rainer Beck / Arthur Utz / Stephen Holloway) • Catalysis • Novel Systems ______(Franz Geiger / David King / (Andy Tsou / Kenji Urayama / Berit Hinnemann / Bengt Kasemo) Tomiki Ikeda / B.R. Ratna) • Liquid-Liquid Interfaces • Biomolecular Networks and Biological ENGINEERING SCIENCES FOR (Heather Allen / Robert Walker / Systems (1) SPACE EXPLORATION Ilan Benjamin) (John Enns / Lynden Archer / Julie Kornfield / LES DIABLERETS CONFERENCE CENTER • Dynamics of Adsorption Nathan Ravi / Ferenc Horkay) LES DIABLERETS, SWITZERLAND (Eckart Hasselbrink / Axel Gross / • Poster Sessions AUG 21-26, 2005 Mary Jane Shultz / John Morris) (Henning Winter / Manika Varma-Nair) G. PAUL NEITZEL, CHAIR • Gas-Liquid Interfaces • Physics of Gels JAMES WARREN, VICE CHAIR (Akihiro Morita / Pavel Jungwirth / (Athene Donald / Bela Joos / Gunther Andersson) Murugappan Muthukumar / Christian Holm) • Granular Media & Particulate Flow I • Motions at Surfaces • Theory and Simulation (Pierre Evesque / James Jenkins / (Bret Jackson / Jascha Repp / (Michael Rubinstein / Sergey Panyukov / John Christine Hrenya) Ellen Williams / William Allison) Curro / Alex Levine / • Granular Media & Particulate Flow II • Electron Dynamics Fernando Escobedo) (Iwan Alexander / Masami Nakagawa / Jeffrey (Jane Hinch / Martin Wolf / Giacinto Scoles) • Biomolecular Networks and Biological Morris / Michel Louge) • Reactions at Surfaces Systems (2) • Multiphase Flow (Bruce Garrett / Sylvia Ceyer / (Marie-France Vallat / David Mooney / (Vemuri Balakotaiah / Mark McCready / Matthias Scheffler / Robert Hamers) Dennis Discher / Jian Ping Gong) Andrea Prosperetti) • Young Investigator Presentations • Pot Pourri (1) • Multiphase Heat Transfer (Greg Sitz) (Judit Puskas / Annelise Barron / (Alcina Mendes / Cila Herman / Jan Genzer / Darrin Pochan) Kambiz Vafai / Johannes Straub) ______• Pot Pourri (2) • Nanofluidics (Lynn Loo / Tatiana Budtova / (Marc Smith / Flavio Noca / Joel Plawsky) Chris Robertson / Jennifer West) • Biological Life Support Systems • Evolving Networks (Jitendra Joshi / Katherine Banks / ELASTIN & ELASTIC FIBERS (Francois Lequeux / Luis Dorfmann / Andrew Jackson / Dani Or) KIMBALL UNION ACADEMY Alan Wineman / Will Mars) • Environmental Monitoring MERIDEN, NH • Filled Systems and Nanocomposites (Gary Sayler / Marc Porter / James Lambert) JUL 31-AUG 5, 2005 (Chris White / Jose Kenny / • Applied / Heat Transfer CAY KIELTY, CHAIR Helene Montes / Alan Gent) (Richard Lueptow / Ward TeGrotenhuis / ELAINE DAVIS, VICE CHAIR • Special Topic Mark Weislogel / Jamal Yagoobi) (Greg McKenna / Gerald Pollack) • Cryogenics • Structures of Elastic Fiber Molecules (Daniel Beysens / Louis Salerno / (Penny Handford / Fred Keeley / Mohammad Kassemi) John Parkinson / Kristin Kumashiro / ______Tony Tamburro / Clair Baldock / Sacha Jensen) ______• Microfibril and Elastic Fiber Assembly (Brenda Rongish / Dieter Reinhardt / Cay Kielty / Tom Wight / Andras Czirok / Sarah Dallas / Jessica Wagenseil)

visit the frontiers of science: www.grc.org ENZYMES, COENZYMES & • Looking Forward • Development and Turnover of METABOLIC PATHWAYS (special consideration will be given throughout Excitatory Synapses KIMBALL UNION ACADEMY all sessions to include new topics, puzzling (Mike Ehlers / Kim McAllister / MERIDEN, NH observations, and mysterious phenomena) Ann Marie Craig / Monica di Luca / JUL 17-22, 2005 Gary Westbrook) EUGENE MUELLER & ______• Glutamate Receptor Modulation MARTIN TANNER, CO-CHAIRS (Juan Lerma / Suzanne Zukin / SUSAN MILLER & Christoph Mulle / Laurent Fagni) JOHN RICHARD, CO-VICE CHAIRS EVOLUTIONARY & ECOLOGICAL FUNCTIONAL GENOMICS ______• Emerging Systems and Methods THE QUEEN’S COLLEGE (Francisco Wilson / Wilfred A. van der Donk / OXFORD, UK Karen S. Anderson / Homme W. Hellinga / JUL 31-AUG 5, 2005 FERTILIZATION & Joseph A. Krzycki) THOMAS MITCHELL-OLDS, CHAIR ACTIVATION OF DEVELOPMENT • Enzyme Mechanisms I GREG WRAY, VICE CHAIR HOLDERNESS SCHOOL (Nigel G.J. Richards / PLYMOUTH, NH Debra Dunaway-Mariano / • Current Progress and Future Directions JUL 17-22, 2005 Peter J. Tonge / Stewart L. Fisher) (Thomas Mitchell-Olds / / RICHARD CARDULLO, CHAIR • RNA: Enzyme and Substrate / Paul Brakefield) GEORGE GERTON, VICE CHAIR (Barry S. Cooperman / • Genomes and the Environment Tamara L. Hendrickson / Dirk Iwata-Reuyl) (Joy Bergelson / Laurent Keller / • Bioinformatic, Genomic, and Proteomic • Carbohydrates Katie Peichel / Tom Whittham) Approaches to Fertilization and (Gideon J. Davies / Paul J. Berti / • Genomics of Complex Traits Development James H. Naismith / Vern L. Schramm) (Trudy MacKay / Scott Edwards / (Anna Neill / Tim Kroft) • Cofactors Jeff Feder / Bill Jeffery) • Sperm Motility and (Adelbert Bacher / Ruma V. Bannerjee / • Transcription and Evolution (George Witman / Winfield Sale / Holly R. Ellis / Dennis R. Dean) (Dan Hartl / Justin Fay / Carol Lee / Chris Wood / Marc Spehr) • Enzyme Mechanisms II Patricia Wittkopp) • Gamete Maturation (Michael A. Marletta / Johannes Rudolph / • Natural Selection and the Genome (Laurinda Jaffe / Lisa Mehlman / Kenny K. Wong) (Joe Thornton / Dan Hartl / Khaled Machaca) • Frontiers in Enzymology Henrik Kaessman / Sue Wessler) • Functional Membrane Domains in Gametes (Perry A. Frey / Rowena G. Matthews) • Evolutionary Ecology of Microbial (George Gerton / Alexander Travis / Genomes Bart Gadella / Nonunj Tanphaichitr) ______(Anthony Dean / Jennifer Hughes / • Molecular Aspects of Martin Parniske / Derek Smith) Sperm-Egg Interaction • Plant Model Systems in EEFG (Catherine Thaler / Daniel Hardy / EPIGENETICS (Enrico Coen / Joy Bergelson / Masaru Okabe) HOLDERNESS SCHOOL Caroline Dean / Cynthia Weinig) • Signal Transduction in Sperm PLYMOUTH, NH • Evolution of Protein Function (Harvey Florman / Melissa Jungnickel / AUG 7-12, 2005 (Sue Wessler / Anthony Dean / Benjamin Kaupp / Dejian Ren) JUDITH BENDER & Joe Thornton / Mariana Wolfner) • Keynote Lecture CHAO-TING WU, CO-CHAIRS (Anne McLaren) ANNE FERGUSON-SMITH & ______• Signaling Events During Egg Activation STEVEN JACOBSEN, CO-VICE CHAIRS (Jay Baltz / Guillaume Halet / Rafael Fissore / Janice Evans) • Inheritance: Remembrance of Things Past • Gene Activation During Early Development EXCITATORY AMINO ACIDS & (Marisa Bartolomei / Vicki Chandler / (Kathleen Foltz / Michelle Roux / BRAIN FUNCTION Robert Pruitt / James Sherley / others) Fabio Piano / Michael Stitzel) CENTRE PAUL LANGEVIN • Crosstalk: Development and Defense AUSSOIS, FRANCE ( / Jeannie Lee / SEP 4-9, 2005 ______Robert Metzenberg / Benjamin Normark / MORGAN SHENG, CHAIR Amy Pasquinelli / Carmen Sapienza / others) ROBERT MALENKA, VICE CHAIR • Change and the Environment FLORAL & VEGETATIVE VOLATILES (Jean Finnegan / Eric Richards / • Keynote Lectures THE QUEEN’S COLLEGE Suzanne Rutherford / (Susumu Tonegawa / Rob Malenka) OXFORD, UK Emma Whitelaw / others) • High Resolution Studies of Glutamate SEP 11-16, 2005 • Crosstalk: Getting the Message Across Receptors JOHN PICKETT & BRIAN SMITH, CO-CHAIRS (Denise Barlow / James Birchler / (Daniel Choquet / Haruo Kasai / WITTKO FRANCKE & William Kelly / Marjori Matzke / Eric Gouaux / Kristen Harris / ERAN PICHERSKY, CO-VICE CHAIRS Craig Pikaard / Wolf Reik / others) Guosong Liu / Scott Thompson) • One Scoop or Two: Dosage and Gene • Controversies in AMPA • Analytical Techniques for Analysis of Floral Regulation Receptor Trafficking and Vegetative Volatiles (Ueli Grossniklaus / / (Rick Huganir / Roberto Malinow / • Neural Representation of Odors and the Jennifer Marshall-Graves / David Bredt / Ed Ziff) Statistics of Natural Odor Scenes Ortrun Mittelsten Scheid / others) • Synaptic Plasticity Mechanisms • Biology of Defensive Volatiles • Switching: (Graham Collingridge / Roger Nicoll / • Biology of Attractant Volatiles Decisions, Decisions, Decisions Gina Turrigiano / Yu Tian Wang / • Natural History of Volatiles (Steve Henikoff / / Julie Kauer / John Isaac) Amar Klar / Erika Matunis / Barbara Panning / • Genetic Approaches to Glutamate Jasper Rine / others) ______Receptors and Brain Function I • Everything in its Place: Genome Organiza- (Masayoshi Mishina / Steve Heinemann / tion Peter Seeburg / Joe Z. Tsien) ( / Anne Ferguson-Smith / • Experience-Dependent Plasticity In Vivo Thomas Jenuwein / (Mark Bear / Takao Hensch / Holly Cline / Robert Martienssen / others) Dan Madison / Dan Feldman) • Keeping the Peace • Genetic Approaches to Glutamate (Timothy Bestor / Sarah Elgin / Receptors and Brain Function II Steve Jacobsen / Eric Selker / others) (Jeremy Henley / Josh Kaplan / Villu Maricq / Stefan J. Sigrist)

visit the frontiers of science: www.grc.org FREE RADICAL REACTIONS • Responses to Replication Blockage HETEROCYCLIC COMPOUNDS HOLDERNESS SCHOOL (Tom Kunkel / John Diffley / Alan Lehmann / SALVE REGINA UNIVERSITY PLYMOUTH, NH Jean Wang) NEWPORT, RI JUL 3-8, 2005 • Double Black Diamonds of Repair: DNA JUL 3-8, 2005 MICHAEL SCHMITTEL, CHAIR Cross-Links and Protein-DNA Cross-Links HUW DAVIES, CHAIR JAMES TANKO, VICE CHAIR (Stephen Lloyd / Peter McHugh / JOHN MACOR, VICE CHAIR Laura Neiderhofer/ Alan D. Andrea) • Radicals in Biology I+II • Damage Inducible Checkpoints and • New Synthetic Strategies (Martin Newcomb / Joan Broderick / Signaling (John Macor / Amir Hoveyda / Barry Trost) Joseph Jarrett / Christian Schöneich / (Tony Carr / Steve Elledge / • Synthesis of Heterocycles Chrys Chatgilialoglu) Rodney Rothstein / Yossi Shilo) (Tom Hoye / Joel Hawkins / Robert Batey) • Synthetic Applications of Nitroxides • DNA Damage In Vivo and Somatic Mutation • Heterocyclic Transformations (Armido Studer / Rebecca Braslau / In Vivo (Michael Van Nieuwehnze / Paul Tordo) (Andrew Collins / Bernd Eppe / Vladimir Gevorgyan) • Radical Ions / Radicals and Metals I+II Vern Walker / Shinya Shibutani) • Biologically Significant Heterocycles (Joseph Dinnocenzo / Mitsuo Sawamoto / • Mitochondria: The Other Site of DNA (Karl Scheidt / Greg Wayne / James Franz / Shunichi Fukuzumi / Damage Louis Jungheim / Melanie Sanford / Jack Norton) (William Copeland / Susan Ledoux / Ute Moll / Dennis Wright) • Mechanisms in Radical Chemistry Takehiko Shibata) • Assembly of Complex Heterocycles (James Tanko / Carl Schiesser / • Metals as Co-Carcinogens (Chulbom Lee / Donald Hertzog / Georg Gescheidt / John Walton) (David Wilson / Andy Kligerman / Timothy Guzi / Mark Lautens) • Radicals in Synthesis Anatoly Zhitkovich / Jim Imlay) • New Applications of Heterocycles (Chrys Chatgilialoglu / Armido Studer / (Helen Blackwell / Jieping Zhu / David Crich / Philippe Renaud) ______Jaume Balsells-Padros / Kevin • Radicals and DNA Burgess / Christine Tarby) (Michael Schmittel / Thomas Carell / • Synthesis and Catalysis Shana Kelley) (Carsten Bolm / Jeffrey Johnston) GLOBAL ASPECTS OF TECHNOLOGY • Cheves Walling Lecture • Organometallic and Heterocyclic Synthesis TRANSFER: BIOTECHNOLOGY (David Crich / Bernd Giese) (Mikund Sibi / Timothy Donohoe / THE QUEEN’S COLLEGE Wendy Young / Richard Larock) OXFORD, UK • Heterocycles and Total Synthesis ______SEP 4-9, 2005 (E.C. Taylor / Dean Toste / Erik Sorensen) JOHN KILAMA, CHAIR RICHARD MAHONEY, VICE CHAIR FUEL CELLS ______BRYANT UNIVERSITY • Defining Technology Transfer: Past, SMITHFIELD, RI Present and Future JUL 17-22, 2005 (Maria Freire / John Kilama / Lita Nelsen / HIGH TEMPERATURE CORROSION KLAUS-DIETER KREUER & Robert Mallett) COLBY-SAWYER COLLEGE THOMAS SCHMIDT, CO-CHAIRS • Biotechnology Policy and Legislation NEW LONDON, NH BRIAN BENICEWICZ & (Robert Horsch / George Atkinson / JUL 24-29, 2005 JEREMY MEYERS, CO-VICE CHAIRS Katsuya Tamai) BRIAN GLEESON, CHAIR • Biotechnology Regulatory and Judiciary W QUADAKKERS, VICE CHAIR • The Frontier (Pauline Newman / Philip Grubb / Tony Bates) (D. Wilkinson / D. Thompsett) • Technology Transfer in Developing No information available at press time. Please • Membranes Countries: Case Studies and Policy check the GRC web site for details. (S. Paddison / B. Pivovar / M. Liu) Analysis • Beyond the Membrane (Hanna Kettler / Tony Heher / ______(J. Newman / W. Petitt) Joachim Oehler / Zhu Chen / Victoria Hale) • Durability/Degradation/Diagnostics • University/Industry/Government Partner- (R. Balliet / E. Roduner / M. Inabe / ships HORMONE ACTION IN H.R. Kunz) (Roy Widdus / Jerry Keusch / DEVELOPMENT & CANCER • (Electro)Catalysis: From Ensembles to Jim K. Muhwezi / Mikyung Yun / MOUNT HOLYOKE COLLEGE Single Atoms Susan Finston) SOUTH HADLEY, MA (G. Lindbergh / M. Koper / E. Savinova / • Global Technology Transfer: Its Impact on JUL 10-15, 2005 U. Heiz / P. Serp) Product Development, Trade, Litigation and MAARTEN BOSLAND, DARCY KELLEY & • PEM Fuel Cells: The Only Choice? Bio-Security CHERYL WALKER, CO-CHAIRS (W. Schnurnberger / S. Haile) (Harvey Bale / Norman Neureiter / SHIUAN CHEN & • Are We on the Right Track? J.D. Richard Wilder / Jorge Amigo Castañeda) PAOLO SASSONE-CORSI, CO-VICE CHAIRS (U. Wagner) • Global Business Market in Biotechnology (Calos Morel / Eric Poincelet / Wendy Taylor) • Non-Steroid Hormone Nuclear Receptor • New Paradigms for Assessing Technology ______Mechanisms Transfer (Paolo Sassone-Corsi / (Keith Maskus / Rafael Rangel-Alda / Bruce Spiegelman / David Moore) Marcel D. Mongeon / Charles A. Gardner / GENETIC TOXICOLOGY • Critical Transitions in Development Usha R. Balakrishnan) COLBY-SAWYER COLLEGE (Donald Brown / Jim Truman / Kevin White / • Impact of Technology Transfer on NEW LONDON, NH Alex Schrieber / Cheryl Sisk) Diagnostics and JUL 31-AUG 5, 2005 • Epigenetic Mechanisms in Hormone Action Pharmaceutical Development BENNETT VAN HOUTEN, CHAIR and Imprinting (Robert Ridley / Gail Cassell / ANTONY CARR, VICE CHAIR (John McLachlan / Gail Prins & Shuk-Mei Ho / Nelson Sewankambo / Kulvinder Singh Saini / David Jarrard / Ken Nephew) Rich Mahoney) • Keynote Talks • Prevention and Therapy of Hormone- (Bennett Van Houten / Samuel Wilson / Related Cancer Peggy Jeggo / Marc Vidal) ______(Ronald Ross) • Genotoxicity of Endogenous Lesions and • Animal Models of Hormone Related Cancer BER and Other Disease (Larry Marnett / Sankar Mitra / (Jim Shull / Myles Brown / Priscilla Furth / John Essignman / Akira Yasui / Margaret Jones) Leona Samson) • New Developments in Excision Repair (Jean-Marc Egly / Caroline Kisker / Kioji Tanaka / Cilla Cooper)

visit the frontiers of science: www.grc.org • Genomic and Non-Genomic Signaling by • Hydrogen Mobility • Inhibitory Synaptic Transmission II Nuclear Hormone Receptors (Alexander Skripov / Klaus-Dieter Kreuer / (Ivan Soltesz / Mu-Ming Poo / Bryndis Birnir / (Rakesh Kumar / Jan-Åke Gustafsson / Gunter Majer) Dimitri Kullman / Jean-Claude Lacaille / Ellis Levin) • Magnetism/Novel Measurement Techniques Kai Kaila) • Genomics, Proteomics, and Metabolomics (Peter Vajda / Igor Goncharenko / Jack Rush / • Inhibition, Networks and Plasticity (Shuk-Mei Ho) Zamir Gavra / Gerhard Krexner) (György Buzsáki / Richard Miles / • Hormone Metabolism • Hydrogen-Storage: Global Views Robert Pearce / Kevin Staley / Yu-Tian Wang) (Shiuan Chen / Rajeshwar Tekmal / (Yuh Fukai / Nobuhiro Kuriyama / • Pathology of Inhibition Jose Russo / Thomas Sutter / Gary Sandrock / Scott Jorgensen) (Stefano Vicini / Sheryl Smith / Bob Brueggemeier) Douglas Coulter / Steven Petrou / • Keynote Speaker ______Hanns Möhler / Carolyn Houser / (Donald Coffey / Gerald Cunha) Yves DeKoninck) • Plenary Lecture II (Hannah Monyer) ______IMMUNOCHEMISTRY & IMMUNOBIOLOGY THE QUEEN’S COLLEGE OXFORD, UK ______HUMAN GENETICS & GENOMICS AUG 7-12, 2005 SALVE REGINA UNIVERSITY GARY KORETZKY, CHAIR NEWPORT, RI , VICE CHAIR INORGANIC CHEMISTRY JUL 24-29, 2005 SALVE REGINA UNIVERSITY ERIC GREEN, CHAIR • B and T Cell Development NEWPORT, RI JAMES LUPSKI, VICE CHAIR (Meinrad Busslinger / Katia Georgopoulos / JUL 17-22, 2005 Rudolf Grosschedl / GEORGE STANLEY, CHAIR • Opening Keynote Juan Carlos Zuniga-Pflucker) BAHRAM MOASSER, VICE CHAIR () • Immune Cell Recognition • Human Haplotype Structure (Mitchell Kronenberg / D. Branch Moody / • Session 1 (David Altshuler / Mark Daly / / Max Cooper) (Bruce Bursten / Jaqueline Kiplinger / Yusuke Nakamura / David Cox / • Innate Immune Responses David Vicic / Paul J. Chirik) / Matt Hurles) (Lewis Lanier / / • Session 2 • Monogenic Disorders Christine Biron / ) (Tracy Hanna / Julia Chan / Janis Louie / (Bob Nussbaum / Steve Warren / • Immune Cell Activation Jonathan Wilker / Dan Mindiola) Raju Kucherlapati / Stuart Orkin) (Doreen Cantrell / Andre Veillette / • Session 3 • Oligogenic Disorders Jurge Tschopp / Warren Leonard) (Jennifer Petoff / Monique Krom / (Nicholas Katsanis / Aravinda Chakravarti / • Memory and Effector Function Mark Mason / Jeffrey Long) Eric Boerwinkle) (Steven Reiner / Rafi Ahmed / • Session 4 • Therapeutics for Genetic Disorders Richard Flavell / Susan Swain) (George Stanley / Roy Periana / (Michael Sereda / Jean Bennet) • Cell Positioning in the Immune System Cassandra Fraser / Pascal Le Floch) • Human Genomics (Jason Cyster / Takashi Nagasawa / • Session 5 (Tom Gingeras / Eric Green / Eddy Rubin / Thomas Boehm / Dan Littman) (William Buhro / Michael D. Fryzuk / Evan Eichler / David Page) • In Vivo Imaging of Immune Responses Christopher Cummins) • Genome Dynamics (Ronald Germain / Matthew Krummel / • Session 6 (Nigel Carter / John Moran / Mike Wigler / Sebastian Amigorena / Ellen Robey) (Andrew Maverick / William Geiger / Jim Sikela / Jim Lupski) • Host/Pathogen Interactions Makoto Fujita / Peter Caravan) • Chromosome Biology (Anne O’Garra / Jorge Galan / • Session 7 (Hunt Willard / Wendy Bickmore / / Emil Unanue) (Kim Dunbar / Karl Wieghardt / Molly Przeworski) • Termination of Immune Responses Susan Kauzlarich / Stephen Koch) • Closing Keynote (Fiona Powrie / Alexander Rudensky / • Session 8 () Arlene Sharpe / Michael Lenardo) (Jim Mayer / Russell Hughes / Francis DiSalvo / David Tyler / ______Clifford Kubiak) • Session 9 (Bahram Moasser / Richard Eisenberg / Harry Gray) HYDROGEN-METAL SYSTEMS INHIBITION IN THE CNS COLBY COLLEGE COLBY-SAWYER COLLEGE WATERVILLE, ME NEW LONDON, NH ______JUL 10-15, 2005 JUL 3-8, 2005 RONALD GRIESSEN & ISTVAN MODY, CHAIR TERRENCE UDOVIC, CO-CHAIRS CHRIS MCBAIN, VICE CHAIR INORGANIC GEOCHEMISTRY CRAIG JENSEN & PROCTOR ACADEMY KLAUS YVON, CO-VICE CHAIRS • Plenary Lecture I ANDOVER, NH () JUL 31-AUG 5, 2005 • Destabilized Hydrides/Imides • Anatomy and Physiology of Interneurons I JEAN CLINE & STEVE GARWIN, CO-CHAIRS (Robert Bowman / John Vajo / Shin-ichi (Tamás Freund / John Rubenstein / CHRISTOPH HEINRICH, VICE CHAIR Orimo) Rosa Cossart / Zoltán Nusser / Alain Marty) • Complex Hydrides • Structure and Function of • Economic Geology: (Volodymyr Yartys / Wojciech Grochala / GABA Receptors I The View from Industry Ragaiy Zidan / Andreas Züttel / (Robert Macdonald / (John Thompson / Greg Hall / Hendrik Brinks) Cynthia Czajkowski / Richard Olsen) Jon Hronsky / Doug Kirwin) • New Hydrogen-Storage Materials • Data Blitz • Global Tectonics, Magmatism and the (Keith Ross / Wendy Mao) (Chris McBain) Generation of Magmatic-Hydrothermal • First-Principles Studies • Structure and Function of Fluids (Mei-Yin Chou / Taner Yildirim / GABA Receptors II (David John / Cornel de Ronde / Michèle Gupta / Miquel Salmeron / (Ruth McKernan / Neil Harrison / Robert Loucks / Yasushi Watanabe) Manos Mavrikakis) Stephen Moss / David Weiss / • Global Tectonic Settings of Ore-Fluid • Hydrides for Batteries Werner Sieghart) Generation in Oceanic and Continental (Annick Percheron-Guégan / • Inhibitory Synaptic Transmission I Basins Peter Notten / Michel Latroche) (Alex Thomson / Massimo Scanziani / (Murray Hitzman / Ross Large / • Thin Films Matt Jones / Peter Jonas / Mark Farrant) Jamie Wilkinson) (Astrid Pundt / Cyril Chacon / Bjorgvin Hjörvarsson / Bernard Dam)

visit the frontiers of science: www.grc.org • Chemical Processes Controlling Fluid and • New Sources and Applications LIQUID CRYSTALS Solute Transport (Tom Settersten / Nils Hansen / COLBY-SAWYER COLLEGE (Phil Candela / Frank P. Bierlein / Robert Walker / Dahv Kliner) NEW LONDON, NH Daniel Core / John Mavrogenes) • Turbulence and Diagnostics Interactions JUN 19-24, 2005 • Physical Factors Driving and Localizing (Andreas Dreizler / Dirk Geyer / OLEG LAVRENTOVICH, CHAIR Fluid Flow Noel Clemens) GREGORY CRAWFORD, VICE CHAIR (Larry Cathles / Larryn Diamond / • Laser Diagnostics of Thomas Driesner & Sebastian Geiger) Biological Phenomena • Biological Aspects • Chemical Traps for Economic Metal (Jurgen Wolfrum / Clemens Kaminski / (Françoise Livolant / Cyrus Safinya / Precipitation Andreas Brockhinke / Houston Miller) Gerard Wong) (Mark Reed / Robert Moritz & K. Kouzmanov / • Particulates • Thin Films and Membranes Brian Rusk, Mark Reed & John Dilles / (Gregory Smallwood / Joe Mauderly / (Sarah Keller / Ka Yee Lee / Yuka Tabe) John Muntean & Jean Cline) Kim Prather) • Sensors and Drugs Delivery • Physics of Fluid Focusing and • Engine Applications (Nicholas Abbott / Joseph Zasadzinski / Hydrothermal Heat Balance (Volker Sick / Jay Jeffries / Jonathan Selinger) (Jeremy Richards / Stephen Cox / Wieland Koban / Scott Sanders) • Self-Assembly and New Phases Raul Madrid) • Diagnostics in Multi-Phase Flows (Noel Clark / Gregory Grason / • Remobilization and Secondary Enrichment (Christof Schulz / Hope Michelsen / Carsten Tschierske) of Metals in the Michael Drake) • Defects and Patterns Supergene Environment • Non-Linear Optics (Maurice Kleman / Robert Kusner) (James Saunders / George Brimhall / (Michael Brown / Bob Lucht / • Theories and Simulations of Steve Enders / Ming-Kuo Lee) Zhungshan Li / Robert Stevens) Molecular Order • Research Frontiers in Hydrothermal • Hot Topics: A Selection From the (Lech Longa / Claudio Zannoni) Processes and Economic Geology Contributed Posters • Colloids and Liquid Crystals (Antonio Arribas / Nick Archibald / Robert (Paul Ewart) (Igor Musevich / Seth Fraden) Bodnar / Steve Kesler / Richard Sillitoe / • Liquid Crystalline Elastomeres Moira Smith) ______(Patrick Mather / Robert Meyer) • Optics and Electrooptics ______(Diederik Wiersma / Dirk Broer) LIPIDS, MOLECULAR & CELLULAR BIOLOGY OF ______INTERIOR OF THE EARTH KIMBALL UNION ACADEMY MOUNT HOLYOKE COLLEGE MERIDEN, NH SOUTH HADLEY, MA JUL 24-29, 2005 LIQUIDS, CHEMISTRY & PHYSICS OF JUN 12-17, 2005 JEAN VANCE, CHAIR HOLDERNESS SCHOOL LARS STIXRUDE, CHAIR CHARLES MARTIN, VICE CHAIR PLYMOUTH, NH GORAN EKSTROM, VICE CHAIR JUL 24-29, 2005 • Mechanisms of Lipid Homeostasis GRAHAM FLEMING, CHAIR • Physics and Chemistry of the Core-Mantle (William Dowhan / Dennis Vance / DAVID REICHMAN, VICE CHAIR Boundary Joseph Goldstein / Michael Brown) (Thorne Lay / Don Helmberger / Kei Hirose) • Cholesterol Metabolism • Complex Fluids • Structure and Evolution of the Core (Laura Liscum / David Russell / (David Grier / Mike Cates) (Bruce Buffett / Dion Heinz / Miaki Ishii) Jerome Strauss / Randy Hampton / • Quantum Processes in Liquids • Geomagnetic Field and the Geodynamo Russell DeBose-Boyd) (Eran Rabani / Brigitte Whaley / Nancy Makri) (Cathy Constable / Andrew Jackson / • Interorganelle Lipid Transport • Ionic Liquids Johannes Wicht) (Dennis Voelker / Masahiro Nishijima / (Paul Madden / C. Austen Angell) • Temperature and Composition of the Lower Richard Pagano / Gerrit van Meer) • Glass Transition Mantle • Phospholipid Metabolism (Nathan Israeloff / Juan P. Garrahan / (Guy Masters / Guillaume Fiquet / (Suzanne Jackowski / Howard Goldlfine / Ranko Richert) Guust Nolet / Paul Tackley) Rosemary Cornell / Neale Ridgway / • Confined Liquids • The Transition Zone Arnold Strauss) (Steve Granick / Rustem Ismagilov) (Barbara Romanowicz / Daniel Frost / • Derivatives of Fatty Acids • Liquids/Biological Interfaces & Systems Jeroen Ritsema) (Edward Dennis / Thomas McIntyre / (L. Mahadevan / Ka Yee C. Lee / • Plate Tectonics and Deep Mantle Structure William Smith) Gerard Wong) (Michael Manga / John Lassiter / • Fatty Acid Biosynthesis • Equilibrium Properties from Adrian Lenardic / Shijie Zhong) (Chuck Rock / John Cronan / Non-Equilibrium Experiments • Planetary Interiors Diego de Mendoza / David Bernlohr / (Gerhard Hummer / Jan Liphardt) (Catherine Johnson / Marc Parmentier / James Ntambi) • Nonlinear Spectroscopic Studies of Liquid Sabine Stanley) • ABC Transporters and Transmembrane Dynamics • Evolution of the Mantle Lipid Flux (Robin Hochstrasser / Phillip Geissler / (Erik Hauri / Paul Asimow / (Chris Raetz / Peter Edwards / Martin Zanni) Al Hofmann / Don Porcelli) Chip Davis / ) • Final Presentation • Unresolved Problems • Triacylglycerol Metabolism, (Robert Zwanzig) (Alex Halliday / Bernard Wood / Obesity and Diabetes David Stevenson) (Rosalind Coleman / Dawn Brasaemle / ______Karen Reue / Hei-Sook Sul / Chris Newgard) ______• Regulation of Lipid Metabolism (George Carman / Bruce Spiegelman / MAGNETIC RESONANCE Carl Sparrow / Charles Martin) CONNECTICUT COLLEGE LASER DIAGNOSTICS IN COMBUSTION NEW LONDON, CT MOUNT HOLYOKE COLLEGE ______JUN 5-10, 2005 SOUTH HADLEY, MA MICHAEL MEHRING, CHAIR JUL 31-AUG 5, 2005 SHIMON VEGA, VICE CHAIR MARK ALLEN, CHAIR PAUL EWART, VICE CHAIR • From Physics to Medicine (Michael Mehring / / Kamil • Advances in Imaging Techniques Ugurbil) (Campbell Carter / Jonathan Frank / • Structural Investigations with Alaa Omrane) Pulsed NMR and EPR (Kamil Ugurbil / Jack Freed / Beat Meier / Tatyana Polenova)

visit the frontiers of science: www.grc.org • External and Remote Detection • Epithelial Mesenchymal Interactions in MECHANISMS OF CELL SIGNALLING (Alex Pines / Bernhard Blümich / Breast Cancer HONG KONG UNIVERSITY OF Song I-Han / Yung-Ya Lin) (Terri Wood / Lewis Chodosh / Claire Lewis / SCIENCE & TECHNOLOGY • From Strongly Correlated Solids to John Condeelis / Derek Radisky) HONG KONG, CHINA Proteins • Developmental Transitions I JUN 12-17, 2005 (Robert G. Griffin / Claude Berthier / (Charles Streuli / Lothar Hennighausen / SHUH NARUMIYA, CHAIR Jürgen Haase / Kurt Zilm / David Weliky) Anthony Brown / Caroline Alexander) JONATHAN CHERNOFF, VICE CHAIR • High Frequency and Complex Pulse • Developmental Transitions II Sequences for Structure Determination in (Pam Cowen / Robert Rhoads / • Signal Transduction and Cytoskeletal Biological Systems Christine Watson / Cynthia Zahnow / Dynamics (Beat Meier / Edgar Groenen / Gertraud Robinson) (David Pellman / Mike Rosen / Judith Herzfeld / Steffen J. Glaser) • Sex Steroid Hormone Regulation in Fred Wittinghofer / Tadaomi Takenawa) • Single Electron Spin Detection and Spin Mammary Development and Cancer • Cell Polarity and Migration I Manipulations in Semiconductors (Robert Clarke / Sandy Haslam / ( / Rick Firtel / (Claude Berthier / Dan Rugar / H.W. Jiang / Leena Hilakivi-Clarke / Gabriela Dontu) Alexander van Oudenaarde / Klaus Hahn / Martino Poggio) • Lactation Zhenbiao Yang) • Tales and Novel Concepts of NMR (Margaret Neville / John Wysolmersky / • Cell Polarity and Migration II (Warren Warren / / Jim McManaman / Sarah Berry / (Greg Gundersen / Fred Chang / Jonathan Jones / Lucio Frydman) Christopher Linington) Tetsu Akiyama / Atsuko Kodama) • From Microtesla NMR to EPR/NMR • Peptide Hormone Signaling and Growth • Cell Adhesion Structure-Function Investigations Factors (Yoshimi Takai / Hans Bos / (Kurt Zilm / John Clarke / Robert Tycko / (Paul Kelley / Christopher Ormandy / Martin Schwartz / Valerie Weaver) Tim Cross / Yuri D. Tsetkov) Nelson Horseman / • Cell Proliferation and • BOOMERANG and Mary Helen Barcellos-Hoff / Sacha Howell) Malignant Transformation I Structural Investigations • Cell Cycle, DNA Repair Pathways, (Dafna Bar-Sagi / / (Shimon Vega / Daniel Weitekamp / Chromosomal Abnormalities and David Tuveson / Fuyuhiko Tamanoi) Thomas Prisner / Lyndon Emsley) Breast Cancer • Cell Proliferation and (Jeffrey Rosen / Joe Jerry / Joe Grey / Malignant Transformation II ______Ralph Scully / Elizabeth Musgrove) (Chris Marshall / Yukihiko Kabuyama / • Keynote Speaker John Blenis / Jonathan Chernoff) (Jeffrey W. Pollard / Robert Weinberg) • Axon Guidance and Synaptic Plasticity (Linda van Aelst / Yukiko Goda / MALARIA Mu-Ming Poo / Yuh-Nung Jan / Didier Job) THE QUEEN’S COLLEGE ______• Mitosis and Cytokinesis OXFORD, UK (Tony Hyman / Bruno Goud / AUG 21-26, 2005 Ed Manser / Robert Rottapel) ANDY WATERS, CHAIR MATRIX METALLOPROTEINASES • GTPases in the Immune System KEVIN MARSH, VICE CHAIR BIG SKY RESORT (Doreen Cantrell / Nagahiro Minato / BIG SKY, MT Victor Tybulewicz) • Molecular Aspects of and AUG 28-SEP 2, 2005 Drug Targets ZENA WERB, CHAIR (Nick White / Steve Ward / CARLOS LOPEZ-OTIN, VICE CHAIR ______Michael Ferdig / John Hyde) • Genomes and Post Genome Analyses • The Metalloproteinase Toolbox: Structural (Dan Carucci / Chris Newbold / Jane Carlton / Biology of Enzymes and Substrates, the MECHANISMS OF MEMBRANE TRANSPORT Xinzhuan Su / Ray Hui) Degradome, Visualizing MMPs TILTON SCHOOL • Molecular Aspects of Infection (Zena Werb / Ben Cravatt / Chris Overall / TILTON, NH (David Conway / Stefan Kappe / Ambra Pozzi / Lynn Matrisian) JUN 5-10, 2005 Ute Frevert / Dominique Soldati) • Metalloproteinases in Stem Cells, ROBERT STROUD, CHAIR • Invasion Development and Aging PETER MALONEY, VICE CHAIR (Tony Holder / / (Lynn Matrisian / Judy Campisi / Chetan Chitnis / Masao Yuda / Maria Mota) Andrea Page-McCaw / Kiyoji Nishiwaki / • Principles of Membrane • Transmission from Host to Vector Beate Heissig / Joe Madri / Judith Kimble) Protein Construction (Geoff Targett / Oliver Billker / • Metalloproteinases in Inflammation (Michael Wiener / Don Engelman / Bob Sinden / Ken Vernick) (Dylan Edwards / Ian Clark / Steven White / James Bowie) • Immunity and Malaria Farrah Kheradmand / Rama Khokha) • The Amt/MEP/Rh family and Gas Transport (Eleanor Riley / Jean Langhorne / • Metalloproteinases in the Nervous (Robert Stroud / Shahram Khademi / / Laurent Renia / Chris System-Neurodegenerative Diseases, Connie Westhoff / Walter Boron) Hunter) Memory and Aging • Translocation of Proteins • Gene Expression (Chris Overall / Leszek Kaczmarek / (Michael Wiener / Thomas Rapaport) (Dyann Wirth / Kirk Deitsch / Gary Rosenberg / Kathy Conant / • Channels that are Transporters and Vice Artur Scherf / Henk Stunnenberg) Falk Fahrenholz / Voon Wee Yong) Versa • Molecular Aspects of Pathology • ADAMS and ADAMTS Families in (Chris Miller / Tzyh-Chang Hwang / (Kevin Marsh / Alex Rowe / Pete Bull / Development and Disease David Gadsby / Michael Kavanaugh / Joe Smith / Kasturi Haldar) (Ulla Wewer / Marcos Milla / Carl Blobel) David Clapham) • Parasite • Metalloproteinases in Cancer • ABC Transporters (Brian Greenwood / Richard Carter / (Mikala Egeblad / Agnes Noel / (Phil Thomas / Amy Davidson / Andrew Read / David Sibley) Lisa Coussens) Shimon Schuldiner / Kaspar Locher) • Inhibitors of Metalloproteinases- • Major Facilitators ______TIMPs and Others (Peter Maloney / Manuel Palacin / (Rama Khokha / Vincent Dive / Ron Kaback / Susan Buchanan) Richard Dyer / Caroline Owen) • Regulation of Transporters • Metalloproteinase Gene Regulation, Cell (Robert Edwards / Barouk Kanner / MAMMARY GLAND BIOLOGY Biology Activation and Receptors Cecilia Canessa / David Gadsby / SALVE REGINA UNIVERSITY (Rafi Fridman / Alex Ullrich / Nabil Seidah / Bernd Fakler / Nancy Andrews) NEWPORT, RI Carlos Lopez-Otin / Alicia Arroyo / • Synaptic Transporters JUN 12-17, 2005 Catherine Muller) (Sandy Bajjalieh / Jean-Yves Lapointe / Robert JEFFREY POLLARD, CHAIR • MMPs- The Next Generation Edwards / Eric Gouaux / Susan Amara) ROBERT CLARKE, VICE CHAIR (Carlos Lopez-Otin / Roger Y. Tsien / • P-Type ATPases Chris Overall / Dylan Edwards) (David Stokes / Rhoda Blostein / • Keynote Speaker Barry Rosen / Jose Arguello / (Jeffrey W. Pollard / Tak Mak) ______Svetlana Lutsenko)

visit the frontiers of science: www.grc.org • Mechanisms of Water Transport MICROBIAL POPULATION BIOLOGY MOLECULAR & CELLULAR BIOENERGETICS (Henning Stahlberg / Thomas Walz / PROCTOR ACADEMY KIMBALL UNION ACADEMY William Harries) ANDOVER, NH MERIDEN, NH • Late Breaking News JUL 17-22, 2005 JUN 26-JUL 1, 2005 MARGARET RILEY, CHAIR PATRICIA KANE, CHAIR ______PAUL RAINEY, VICE CHAIR STANLEY DUNN, VICE CHAIR

• How Far Have We Come; The Past Twenty • Respiratory Chain Complexes: Structure, Years of Microbial Population Biology Function, and Mechanism MECHANOTRANSDUCTION & GRAVITY (Bruce Levin / Dan Hartl / Dan Dykhuizen / (Shelagh Ferguson-Miller / Leo Sazanov / SIGNALING IN BIOLOGICAL SYSTEMS John Roth / Monica Riley) Ulrich Brandt / Edward Berry) THE UNIVERSITY OF NEW ENGLAND • Phage Diversity and Evolution • Bioenergetics and Disease BIDDEFORD, ME (Jim Bull / Roger Hendrix / Holly Wichman / (David Nicholls / Eric Schon / Martin Brand / JUL 24-29, 2005 Susanna Twiddy) Daniel Ricquier / Valina Dawson) , CHAIR • Protist Diversity and Evolution • Metal Ion Homeostasis and PAUL BLOUNT & (Graham Bell / / Laura Katz) Mitochondrial Function GLORIA MUDAY, CO-VICE CHAIRS • Microbial Ecology (Jerry Kaplan / Sabeeha Merchant) (Colleen Cavanaugh / Sallie Chisolm / • V- and F-type ATPases: Structure, Function, • Mechanosensing in Animals, Plants and Peter Morin / Takema Fukatsu) and Mechanism Microbes • Cooperativity Among Microbes (Brian Cain / Stephan Wilkens / (Ching Kung / Maurice Kernan / (Greg Velicer / Ben Kerr / Ashleigh Griffin) Bettina Bottcher / Nathan Nelson / Ruth Anne Eatock) • Ecology and Evolution of Infectious Masasuke Yoshida) • Cellular and Tissue Response to Mechani- Disease • Sensing and Regulating pH Gradients In cal Stimuli or Gravity (Abigail Saylers / Nicole Perna / Vivo (Simon Gilroy / Jeffrey Dazen / / Carl Bergstrom) (Michael Forgac / Rajini Rao / Fiona Karet / Mark Johnson) • Genome Enabled Studies Raul Martinez-Zaguilan) • Mechanosensitive Channels I (Jeffrey Blanchard / Fred Blattner / Jonathon • Poster Discussion Workshops (Eduardo Perozo / Ian Booth) Eisen / Bernhard Palsson) (L. Shannon Holliday / Terry Krulwich / • Genetic Approaches to Gravity Signaling • Phenotype/Genotype Mapping Diego Gonzalez-Halphen / Robert Gennis) (Elizabeth Haswell / Jiri Friml) (Dom Schneider / Rob Dorit / Rich Lenski) • Biosynthesis and Assembly of Respiratory • Mechanical and Gravity • Evolution of Sex Chain Complexes Signal Transduction (Rosemary Redfield / Lin Chao / Sally Otto) (Rosemary Stuart / Antoni Barrientos / (Natasha Raikhel / Angus Murphy / Carlos Moraes / Thomas Langer) Mike Gustin) • Genomic and Proteomic Approaches to • Mechanosensitive Channels II ______Bioenergetics (Miriam Goodman / David Corey / (Andre Goffeau / Douglas Wallace / Klaus Schulten) Volker Muller / Brad Gibson / Brigitte Meunier) • Genetic Approaches to MICROFLUIDICS, PHYSICS & CHEMISTRY OF • New Frontiers in Mitochondria Biology Mechanotransduction MAGDALEN COLLEGE (Peter Pedersen / Ronald Butow / (Teresa Nicolson / Cori Bargmann) OXFORD, UK David Chan / Vamsi Mootha) • Hot Topics in Mechanotransduction and AUG 21-26, 2005 Gravity Signaling LAURIE LOCASCIO, CHAIR ______SABETH VERPOORTE, VICE CHAIR • Single Molecule Manipulation and Measurement MOLECULAR MECHANISMS OF (Sabeth Verpoorte / Yoshinobu Baba / Jean- MICROBIAL ADHESION MEDICINAL CHEMISTRY Loius Viovy) SALVE REGINA UNIVERSITY COLBY-SAWYER COLLEGE • Multiphase Systems NEWPORT, RI NEW LONDON, NH (Steven Haswell / David Weitz / Wyatt AUG 7-12, 2005 AUG 7-12, 2005 Vreeland / Rustem Ismagilov / Kathleen PASCALE COSSART & S. DAVID KIMBALL, CHAIR Stebe) , CO-CHAIRS JOHN LOWE, VICE CHAIR • Nanobiotechnology BONNIE BASSLER & GEORGE HARTMAN, VICE CHAIR ELECT (Luke Lee / Harold Craighead / Paul Cremer) ARTURO ZYCHLINSKY, CO-VICE CHAIRS • Pushing the Boundaries of Separation • Kinase Inhibitors (James Landers / Gert Desmet / • Opening Lectures (Andrew Mortlock / Walter Ward / Annelise Barron / Anup Singh) (Jules Hoffmann / Jeff Gordon) Paul Manley / Michael Kaufman / • Integrated Systems • Structure/Localization Brian Werneberg) (Brian Kirby / Richard Mathies / Mark Burns) (Staffan Normark / Maria Sandkvist / • The Hit-to-Lead Process • Nano and Microscale Transport Christine Jacobs / Eduardo Groisman / (Amy Ripka / Samuel Gerritz / Brian Shoichet / (Gary Slater / Hans Hermann Gerdes / Albert Guy Cornelis) Bill Michne / Dennis Underwood) van den Berg / Don DeVoe / Sandra Troian) • Microbe-Plant Interactions • Biomarkers and PET Imaging • Crossover (Staffan Normark / / (Balu Balusubramanian / Chet Mathis / (Paul Yager / Olga Vinogradova / David Grier) Sharon Long / Jeff Dangl) Doug Dischino / Richard Hargreaves) • Cells in Microfluidics • Multicellularity and Biofilms • HCV Antivirals (Peter Wilding / Daniel Chiu / Minoru Seki / (Philippe Sansonetti / Ute Römling / (Michael Bos / Ralf Bartenschlager / Jon Cooper / Sangeeta Bhatia) Iñigo Lasa / George O’Toole / Robert Perni / Pierre Beaulieu / Dick Storer) • Nanofluidics versus Microfluidics Jean-Marc Ghigo) • New Directions in Antibiotics (Laurie Locascio / Andreas Manz / Jan Eijkel) • Immunity (John Primeau / Paul Aristoff / (Philippe Sansonetti / Arturo Zychlinsky / Dean Stamos / Zhengyu Yuan) Stephen Girardin / Michael Starnbach) • Protease Inhibitors for Treating Pulmonary ______• Eukarya-Bacteria Signaling Inflammatory Disorders (Brett Finlay / Dan Portnoy / Deborah Hogan / (Bruce Maryanoff / Clive Page / Norma Andrews / Javier Pizarro-Cerda) Michael Greco / Zhongli Gao / John Link) • Toxins and Virulence Factors • New Treatments for Ophthalmological (Brett Finlay / Gisou van der Goot / Disorders Emmanuel Lemichez / Michael Caparon) (Peter Klimko / Jun Inoue / • Intra- and Extra- Cellular Signaling Yongxin Han / Jesse May) (Jorge Galán / Richard Losick / • Special Topics Bonnie Bassler / Gisela Storz / (Mary Mader) Diego de Mendoza) • Chair’s Talk (Roy Vagelos)

visit the frontiers of science: www.grc.org • Pneumococcus-Streptococcus • Exploiting Hypoxia in Cancer Treatment • Unusual Cytoskeletons and Host-Pathogen (Jorge Galán / Birgitta Henriques / (J. Martin Brown / Adrian Harris / Interactions Paul Sullam) Giovanni Mellilo / William Wilson) (Pascale Cossart / David Sibley / • Hip-Hop on : Cytoskeleton Isabelle Tardieux / Matt Welch) ______Takes Center Stage in Cancer Therapy • Cytoskeleton and Cell Signaling (Paraskevi (Evi) Giannakakou / Ken Downing / (Gregg Gundersen / Shuh Narumiya / Paraskevi Giannakakou / Tito Fojo / Klaus Hahn / Pierre Gönczy) Gregg Gundersen) MOLECULAR MEMBRANE BIOLOGY • Unconventional Therapeutic Chemical PROCTOR ACADEMY ______Approaches ANDOVER, NH (Greg Verdine / Greg Verdine / Julian Adam / JUL 10-15, 2005 Saul Rosenberg) SANDRA SCHMID, CHAIR MULTI-DRUG EFFLUX SYSTEMS BENJAMIN GLICK, VICE CHAIR MAGDALEN COLLEGE ______OXFORD, UK • Membrane Fusion AUG 28-SEP 2, 2005 (Bill Wickner / Ed Chapman / ATTILIO DI PIETRO & Reinhard Jahn / Margaret Kielian) MOLYBDENUM & TUNGSTEN ENZYMES LAURA PIDDOCK, CO-CHAIRS • Diverse Translocation Machineries THE QUEEN’S COLLEGE RICHARD BRENNAN & (Tom Rapoport/ Hidde Ploegh / OXFORD, UK SUSAN COLE, CO-VICE CHAIRS Nikolaus Pfanner / Rick Rachubinski / JUL 10-15, 2005 Art Johnson) MICHAEL JOHNSON & • Keynote Session - Multi-Drug Resistance • Intracellular Sorting Mechanisms TRACY PALMER, CO-CHAIRS Overview: From Bacteria to Man ( / Shou-ou Shan / CAROLINE KISKER & (Laura Piddock / Attilio Di Pietro / Hugh Pelham / Linda Hicke) ALASTAIR MCEWAN, CO-VICE CHAIRS Hiroshi Nikaido / ) • Organizing Membrane Subdomains • Biological Significance of Multi-Drug Efflux (Ira Mellman / Rob Parton / Matthias Weiss / • Keynote Speakers Pumps Catherine Rabouille) (K.V. Rajagopalan / Les Dutton) (Marc Ouellette / Hendrik Van Veen / • Membrane Bending/Fission • Molybdenum Biosynthesis Francisco Gamarro / Herman Koepsell) (Janet Shaw / Alberto Luini / (Ralf Mendel / Guenter Schwarz / • Molecular Basis of Drug Recognition and Harvey McMahon / Michael Kozlov) Petra Haenzelmann / Herman Schindelin / Transport • Conformational Dynamics at the Silke Leimkuhler / Emilio Fernandez) (Susan Cole / Melissa Brown / Membrane • Sulfite Oxidases Anne H. Dantzig / Balazs Sarkadi) (Bill Balch / Don Engelman / (Caroline Kisker / John Enemark / Sue Bailey / • Structure and Molecular Mechanism of David W. Andrews / Ehud Isacoff / Partha Basu) Efflux Pumps Diane Papazian) • DMSO Reductase Family (Arnaud Ducruix / Taiji Nakae / • Lipids and Compartment Identity (Al McEwan / Fraser Armstrong / Geoffrey Chang / Anthony M. George) (Pietro DeCamilli / Jean Gruenberg / Chantal Iobbi-Nivol / Dick Holm) • Mechanism of Energy Supply and Coupling Temo Kurzchalia / Scott Emr) • Nitrate Reductases to Drug Efflux • Organelle Biogenesis and Dynamics (David Richardson / Joel Weiner / (Frances Sharom / Eitan Bibi / (Graham Warren / Jodi Nunnari / Katrin Fischer / Simon De Vries / Jose Moura) Jean-Michel Jault / Christopher Higgins) Jennifer Lippincott-Schwartz / / • Xanthine Oxidase Family • Regulation of Efflux Pumps Judith Klumperman) (Russ Hille / Takeshi Nishino / Florian Bittner / (André Goffeau / Richard Brennan / • Transport Carrier Formation Matthias Boll / Marty Kirk / Charlie Young) Scott W. Moye-Rowley / Katleen Scotto) ( / Bruno Antonny / • Tungsten Enzymes • Clinical Relevance Juan Bonifacino / Vivek Malhotra) (Dave Garner / Mike Adams / (Susan Bates / Glenn Kaatz / Bill Shafer / Peter Kroneck / Maria Romao) Antonio T. Fojo / Maria Pia DePasquale) ______• Applied, Environmental and Medical • Hot Topics/Controversies: Aspects Drug Transport Mechanism (Ed Stiefel / Enrico Garattini / Joanne Santini / (Wil Konings / Kim Lewis) Susanne Fetzner / Dave Kelly) • Hot Topics/Controversies: Can Pumps be MOLECULAR THERAPEUTICS OF CANCER Blocked? Will it be Efficient on MDR? COLBY-SAWYER COLLEGE (David Hooper / Olga Lomovskaya) NEW LONDON, NH ______• Reflections on the Past and Glimpses into JUL 17-22, 2005 the Future: Functional Genomics and YVES POMMIER, CHAIR Pharmacogenomics ALAN EASTMAN, VICE CHAIR MOTILE & CONTRACTILE SYSTEMS (Toshihisa Ishikawa / Estelle Marrer / COLBY-SAWYER COLLEGE Rheinhold Kerb / Michael Gottesman) • Targeting the Genome NEW LONDON, NH (Christian Bailly / Christian Bailly / JUL 10-15, 2005 Laurence Hurley / John Hartley) VLADIMIR GELFAND, CHAIR ______• Translation Initiation Pathways for Cancer MARGARET TITUS, VICE CHAIR Therapy (Peter Houghton / Peter Houghton / • Keynote Speaker MUSCLE: CONTRACTILE PROTEINS Chris Proud / Sara Kozma / Jerry Pelletier) (Joel Rosenbaum) COLBY-SAWYER COLLEGE • c-myc and Transcription • Force Production NEW LONDON, NH (Peter K. Vogt / Peter K. Vogt / (Dan Kiehart / Tom Pollard / Tom Roberts / JUL 3-8, 2005 Edward Prochownik / Lubomir Vassilev / Michael Sheetz) LEE SWEENEY, CHAIR Kevin Gardner) • Centrosome JUSTIN MOLLOY, VICE CHAIR • Tantalizing Thanatos-Cell Death as a (Tim Stearns / Alexey Khodjakov / Therapeutic Goal Steve Doxsey / David Agard) • Overview of the Actomyosin Mechanism (Guido Kroemer / Guido Kroemer / • Cytoskeletal Diseases (James Spudich) John C. Reed / Douglas R. Green / (Erika Holzbaur / Tom Friedman / • Regulation of Myosin Motors and Muscle Craig Thompson) Frédéric Saudou / Kevin Campbell) Contraction • Survival Pathways • Cell Polarity (Kathleen Trybus) (Phillip Dennis / Philip Dennis / (Carole Parent / Rong Li / Ian Macara) • Myosin Movement Beyond the Lever Arm William Pao / Shigeki Miyamoto) • Cell Division (H. Lee Sweeney) • The p53-Mdm2 and Stress Response (Claire Walczak / Kerry Bloom / • Myosin Force and Movement Generation Pathways Don Cleveland / Tarun Kapoor) (Yale Goldman) (Wafik S. El-Deiry / Thanos Halazonetis / • Intracellular Movements • Myosin Force and Movement Andrei Gudkov / Alan Weissman / (Lois Weisman / John Kendrick-Jones / Generation in Muscle Wafik El-Deiry) Jon Scholey / Robert Singer / (Malcolm Irving) David Drubin / John Hammer)

visit the frontiers of science: www.grc.org • Microtubular Motor Function • Protein Chemical Biology • Recognition of Kin and Competitor (Kazuhiro Oiwa) (Suzanne Walker / Ben Davis / (Lori Moreno / Brenda McCowan / • Influence of Load on the Actomyosin Cycle Herbert Waldmann) Olivier Pascalis / Elizabeth Tibbets) of Muscle • Medicinal and Process Chemistry I • From Animals to Robots (Michael Ferenczi) (Ann Weber / Karl Hansen) (Ryohei Kanzaki / Hillel Chiel / Barbara Webb / • Influence of Load on Processive Motors • Medicinal and Process Chemistry II Bijan Pesaran / Charles Higgins / (Claudia Veigel) (Milton Brown / Chris Senanayake) Roy Ritzmann) • Meeting Synthesis • Nutraceuticals and Medicinal Plants • Gene Expression and Behavior (Justin Molloy / Hugh Huxley) (Norman Farnsworth / Qun Yi Zheng / (Henrike Scholz / Charles Whitfield / • Poster Review and Discussion Bruce Lipschutz) Hideako Takeuchi / Adi Mizrahi) (Michael Ostap) • Methodology and Total Synthesis III (Peter Wipf / Eiichi Nakamura) ______• Methodology and Total Synthesis IV (Andy Evans / Paul Wender) NEUROTROPHIC FACTORS MYCOTOXINS & PHYCOTOXINS ______SALVE REGINA UNIVERSITY COLBY COLLEGE NEWPORT, RI WATERVILLE, ME JUN 19-24, 2005 JUN 19-24, 2005 NEURAL CIRCUITS & PLASTICITY BARBARA HEMPSTEAD, CHAIR WAYNE BRYDEN & SALVE REGINA UNIVERSITY DAVID GINTY, VICE CHAIR STEVE MUSSER, CO-CHAIRS NEWPORT, RI KATHLEEN REIN & JUN 26-JUL 1, 2005 • Axonal Control KENNETH VOSS, CO-VICE CHAIRS GINA TURRIGIANO, CHAIR (Nancy Ip / Stephen Strittmatter / Bill Snider / RAFAEL YUSTE, VICE CHAIR Paul Letourneau / Frank Gertler) • Risk Assessment and its Role in • Neurotrophic Factor Signaling I Regulatory Decision Making • Keynote Speaker (Mark Bothwell / Chris Garcia / Phil Barker / (David Miller / Sarah Henry / (Rodolfo Llinas) Sungok Yoon / Carlos Ibanez) Elaine Faustman / Wetzel Gelderblom) • Circuit Formation • Actions in Development I • Toxicogenomics and Molecular Mecha- (Holly Cline / Josh Sanes / Tom Sudhof) (Louis Reichardt / Yves-Alain Barde / nisms of Toxicity • Local Control of Synaptic Properties Perry Bartlett / Shahin Rafii / Luis Parada) (Annie Pfohl-Leszkowicz / Greg Mayor / (Erik Kandel / Kelsey Martin / Mike Ehlers) • Neurotrophic Factor Signaling II Michael Stone / Ed Cleveland / • and Visual Circuitry (Bill Mobley / Marino Zerial / Elisabeth Fisher / Wolfgang Dekant) (Markus Meister / Peter Sterling / Axel Borst) Roz Segal / Simon Halegoua) • Toxin Synthesis and Synthetic Receptors • Olfaction/Gustation • Actions in Adulthood and Disease (Mark Savard / Andrea Bordelais / (Larry Katz / Zach Mainen / Don Katz) (Eric Shooter / Valina Dawson / Robert Gawley / Sergey Piletsky) • CPGs and Spinal Circuits Mark Tuszynski / Holly Ingraham) • Toxin Induction and Biosynthesis (Eve Marder / Ole Kiehn / Nick Spitzer) • Synaptic Actions (Bill Gerwick / Nancy Keller / • Auditory System and (Rita Balice-Gordon / Lino Tessarollo / David Sherman / Cynthia Heil / Vocal Communication Bai Lu / Pietro DeCamlli) Samuel Lo Chun-Lap) (Constance Scharff / Michael Brainard / • Therapeutic Implications • Toxin Mitigation and Control Asif Ghazanfar) (Moses Chao / Freda Miller / Dinah Sah / (Russell Molyneux / Kevin O’Shea / • Persistant Activity and Motor Control Jeff Milbrandt / Lieve Moons) Reinhard Fischer / Anne Desjardins / (Sascha Du Lac / David Tank / • Actions in Development II Greg Doucette) H. Sebastian Seung / Mike Shadlen) (Carol Mason / Susan McConnell / • Detection Technologies for Organisms and • Cortical and Hippocampal Circuitry and Kuo-Fen Lee / Anders Nykjaer) Toxins Plasticity (Ken Voss / Kathryn Coyne / (David McCormick / Massimo Scanziani / ______Kevin James / Rudolf Krska / Chris Taitt) Wendy Suzuki) • Animal Model Studies I (Andrew Gordon / Janet Benson / ______NONLINEAR SCIENCE Raghu Sharma / Roger Coulombe) COLBY COLLEGE • Animal Model Studies II WATERVILLE, ME (Fran Van Dolah / Jerry Rice / NEUROETHOLOGY: JUN 26-JUL 1, 2005 Sven Daenicke / Julie Zaias) BEHAVIOR, EVOLUTION & NEUROBIOLOGY EDWARD OTT, CHAIR • Assessing Chronic Exposure to Toxins MAGDALEN COLLEGE ANNA LIN, VICE CHAIR (Robert MacPhail / Helen Shurz-Rogers) OXFORD, UK AUG 7-12, 2005 • Granular Media ______NICHOLAS STRAUSFELD, CHAIR (Wolfgang Losert / Harry Swinney / CATHERINE CARR & Robert Behringer) PAUL KATZ, CO-VICE CHAIRS • Pattern Formation NATURAL PRODUCTS (Ken Showalter / Leonard Sander / TILTON SCHOOL • Opening Session Hugues Chaté) TILTON, NH (Nicholas Strausfeld / Paul Katz / • Animal Grouping JUL 24-29, 2005 Catharine Carr / Edmund T. Rolls / (Iain Couzin / Chad Topaz) JOSEPH ARMSTRONG, CHAIR Thomas Seeley) • Living Systems FREDERICK LUZZIO, VICE CHAIR • Ecological Neuroethology (Eshel Ben-Jacob / Z. Jane Wang) (Eric Warrant / Anna Gislen / Jochen Zeil / • Fluid Singularities • Total Synthesis Peter Narins / Melissa Bateson / (Tomas Bohr / Detlef Lohse) (Dave Askin / Mark Rizzacasa / Caroly Shumway) • Systems and Singularities Sam Danishefsky) • Evolution of Behavior (Martin Hasler / Shankar Venkataramani) • Catalytic Methodology I (Kathy Rankin / Jonathan Bacon / • Dynamics with Lasers (Tim Jamison / Helene Lebel) Hudson Reeve / Heather Eisthen / (Raj Roy / Thomas Antonsen / Nir Davidson) • Catalytic Methodology II Bernard Crespi / Daniel Robert) • Turbulence (Dan Yang / Huw Davies / Mikiko Sodeoka) • Emergent Behavior (Itamar Procaccia / Anne Juel) • /Biology (Susan Fahrbach / Samuel S. Wang / • Chaos (Bradley Moore / Joern Piel / Dorothey Cheney / Fred Dyer / (Lou Pecora / Edward Lorenz / James Yorke) Nicola Pohl / ) Archero Martinoli) • Methodology and Total Synthesis I • Navigation and Migration ______(Gary Molander / Corine Aubert) (Kathy Gotthard / Anna Gagliardo / • Methodology and Total Synthesis II Thomas Collett / Kenneth Lohmann) (Jim Leighton / Brian Stolz / Erick Carreira)

visit the frontiers of science: www.grc.org NUCLEAR CHEMISTRY • DNA Replication and Recombination • Coordinate Transformations for Action COLBY-SAWYER COLLEGE (Lorena Beese / Phoebe Rice / (Paul Glimcher / Carol L. Colby / NEW LONDON, NH Stephen Benkovic / Karl-Peter Hopfner) Jennifer M. Groh / Terry Stanford / JUN 26-JUL 1, 2005 • DNA Damage, At-Risk Motifs and Repair Luis Populin) PARTHA CHOWDHURY, CHAIR (Samuel Wilson / Cynthia Burrows / • Keynote: Where Do We Go From Here? ROY LACEY, VICE CHAIR Cynthia McMurray) (Lawrence E. Mays / John Maunsell / • Nucleic Acids Structure and Dynamics David L. Sparks) • Advances in Light Nuclei (Carlos Bustamante / Anna Marie Pyle / • Deciding [Where to Look][to Act] • Shell Structure in the 21st Century Vincent Croquette / Michelle Wang) (Jennifer M. Groh / Paul W. Glimcher / Richard • Physics Away from Stability: • Ribosomes and Translation J. Krauzlis / Michele Basso / Neutron-Rich Nuclei (Matthias Hentze / Joseph Puglisi / Paul Gamlin) • Physics Away from Stability: James Williamson / Nenad Ban) • New Approaches / Perspectives on Ocular Proton-Rich Nuclei • Transcription and Motility Disorders • Isomers and Nuclear Structure (Joan Conaway / Karolin Luger / (R. John Leigh / Elizabeth C. Engle / • Symmetries, New and Old Richard Gourse / Jeff Gelles) John S. Stahl / Michael Mustari) • Nuclear Structure and Astrophysics • RNA Processing • Synthesis and Spectroscopy of (Allan Jacobson / Holger Stark / ______Heavy Nuclei Brent Graveley) • Weak Interactions and Nuclear Structure • Exotic Excitation and Decay Modes ______ORGANIC REACTIONS & PROCESSES • Rare Isotope Beams and Techniques BRYANT UNIVERSITY SMITHFIELD, RI ______NUCLEOSIDES, NUCLEOTIDES & JUL 31-AUG 5, 2005 OLIGONUCLEOTIDES STEVEN KING, CHAIR SALVE REGINA UNIVERSITY MATT MCINTOSH, VICE CHAIR NUCLEAR PHYSICS NEWPORT, RI BATES COLLEGE JUN 26-JUL 1, 2005 • New Reactions in Organic Synthesis 1 LEWISTON, ME DONNA SHEWACH, CHAIR (Anna Mapp / Ed Vedejs / David Barnes / JUL 10-15, 2005 JYOTI CHATTOPADHYAYA, VICE CHAIR Sergey Kozmin) THOMAS LUDLAM, CHAIR • Organo Metallic Chemistry in Organic MICHAEL RAMSEY-MUSOLF, VICE CHAIR • Keynote Speaker Synthesis 1 (William Plunkett) (David Collum / Viresh Rawal / Jerry Murry / • Quark Structure of Hadrons and Nuclei • G-quadruplex Interactions Dali Sames / Shannon Stahl) (Barry Holstein / Tony Thomas / (Laurence Hurley / Stephen Neidle / • Recent Developments in Catalysis Paul Eugenio / Latifa Elouadrhiri) Laurence Hurley) (Margaret Hsu / David MacMillan / Sally Gut / • The Spin Structure of the Nucleon • DNA Replication and Fidelity Brian Stolz) (Werner Vogelsang / Lara De Nardo / (Thomas Kunkel / Joann Sweasy / • New Reactions in Organic Synthesis 2 Zein-Eddine Meziani / Steve Vigdor) Floyd Romesberg / Michael Famulok) (Karl Scheidt / Shu Kobayashi / • Phases of QCD Matter I • Innovative Nucleic Acid Tools Jonathan Ellman / Kai Rossen / (Barbara Jacak / Ulrich Heinz / (Anna Khvorova / Oliver Seitz) Tom Rovis / Greg Dake) John Harris / Larry McLerran) • Novel Nucleoside Analogs • Mechanistic Chemistry 1 • Phases of QCD Matter II (Frank Seela / Mitsuo Sekine) (Rodney Parsons / Donna Blackmond / (Frithjof Karsch / Xin-Nian Wang / • Nucleosides in Gene Transfer Therapy Ed Delany) Paul Sorensen / Barbara Jacak) (Margaret Black / Barry Stoddard / • Selectivity and Target Oriented Synthesis • The Structure of Hadrons Svend Freytag) (Lisa Reeder / Steven Burke / (Betsy Beise / Rolf Ent / Barry Holstein) • New Antivirals Scott Rychnovsky / Bernhard Breit / • Nuclear Structure at the Frontier (Karen Anderson) Melanie Sanford) (Rick Casten / Rick Casten / • Hot Topics • Organo Metallic Chemistry in Organic Witek Nazarewicz / Thomas Glasmacher / Synthesis 2 Guy Savard) ______(Dick Schrock / Wayne Luke) • Nuclear Astrophysics • Mechanistic Chemistry 2 (Micael Wiescher / Tony Mezzacappa / (Matt MacIntosh / Daniel Singleton / Madappa Prakash) Guy Lloyd-Jones / Michel Journet) OCULOMOTOR SYSTEM BIOLOGY • The Universe Now • Special Lecture BATES COLLEGE (Larry McLaran / Mark Trodden) (Bill Bailey / Reinhard Hoffmann) LEWISTON, ME • Nuclear Physics and the Standard Model JUN 26-JUL 1, 2005 (Michael Ramsey-Musolf / Joshua Klein / LAWRENCE MAYS & ______Brad Filippone / Paul Souder / Kevin Lesko) JOHN PORTER, CO-CHAIRS NEERAJ GANDHI & ______JENNIFER GROH, CO-VICE CHAIRS ORGANIC THIN FILMS SALVE REGINA UNIVERSITY • Pulling Together the Disparate Parts of Eye NEWPORT, RI NUCLEIC ACIDS Movement Control Systems JUL 10-15, 2005 SALVE REGINA UNIVERSITY (David L. Sparks / Joseph L. Demer / NICHOLAS ABBOTT & NEWPORT, RI Stephen Lisberger / David Zee) MILAN MRKSICH, CO-CHAIRS JUN 5-10, 2005 • Integrative Biology of Extraocular Muscle MARTIEN COHEN STUART & MARTHA FEDOR & JOHN TAINER, CO-CHAIRS (John D. Porter / Linda McLoon / NICHOLAS KOTOV, CO-VICE CHAIRS STEPHEN KOWALCZYKOWSKI & Jean Buttner-Ennever / Stephen J. Goldberg / SCOTT STROBEL, CO-VICE CHAIRS Henry J. Kaminski) • Monolayers • Saccade-Vergence Interaction (David Reinhoudt / Christopher Chidsey / • Chromosome Structure and Maintenance (David S. Zee / Claudio Busettini / Jan Genzer) ( / Elizabeth Blackburn / Michael King / R. John Leigh / • Thin Films Martin Gorovsky) Kathleen Cullen) (Tom Russell / Christine Luscombe / • RNAi and MicroRNA • Integration of Oculomotor Subsubstems Paul Nealey / Jim Watkins) ( / Phillip Zamore / (Kathleen Cullen / J. Douglas Crawford / • Liquid Crystalline Films Leemor Joshua-Tor) Edward G. Freedman / Dora E. Angelaki / (Philippe Poulin / Peter Palffy-Muhory / • RNA-Mediated Regulation in Bacteria Neeraj Gandhi) Timothy Bunning) ( / Robert Batey / • Mechanisms of Sensorimotor Integration • Electrooptical Thin Films Karen Wassarman / Andrew Feig) (Neeraj Gandhi / William C. Hall / (Mary Galvin / Stephen Forrest) Edward L. Keller / John Van Opstal / Raymon M. Glantz)

visit the frontiers of science: www.grc.org • Single Biomolecule Force Measurements PHAGOCYTES PHOTOCHEMISTRY (Yves Dufrene / Deborah Leckband) CONNECTICUT COLLEGE BRYANT UNIVERSITY • Biomolecular Interfaces NEW LONDON, CT SMITHFIELD, RI (Horst Vogel / Ka-Yee Lee / Christopher Lowe) JUN 12-17, 2005 JUL 10-15, 2005 • Scaffolds for Cells MARY DINAUER, CHAIR VAIDHYANATHA RAMAMURTHY & (Kevin Healy / David Mooney / JOEL SWANSON, VICE CHAIR KIRK SCHANZE, CO-CHAIRS Philip Messersmith) LINDA JOHNSTON, VICE CHAIR • Zanvil Cohn Lecture ______(Sam Silverstein) • Molecular, Supramolecular and Chiral • Regulation of Myelopoiesis Photochemistry (Robert Clark / Stu Orkin / Dan Tenen / (N.J. Turro / M. Garcia-Gariba / J.C. Scaiano / Dan Link / Klaus Ley) T. Bach / B. Feringa / R.G. Weiss) ORGANOMETALLIC CHEMISTRY • Actin and Membrane Dynamics • Materials Photochemistry SALVE REGINA UNIVERSITY (Tom Stossel / Fred Maxfield / (L. Brus / G. Calzaferi / NEWPORT, RI Fred Southwick / Becky Worthylake / Anna Köhler / O. Nalamasu) JUL 10-15, 2005 Tom Stossel) • Bio-Photonics and Sensors PAUL FAGAN, CHAIR • Inflammation and Disease (D.G. Whitten / W. Tan / L. Tolbert / P. Ogilby) GERARD PARKIN, VICE CHAIR (Dirk Roos / Dan Kastner / • Energy Conversion Lisa Coussens / John Harlan) (E. Galoppini / H. Arakawa) • Alexander M. Cruickshank Lecturer • Leukocyte/Endothelial Cell Interactions • Physical and Theoretical Photochemistry (Robert Grubbs) (Gary Bokoch / Claire Doerschuk / (B. Schwartz / J. Michl / M. Robb / • Organometallics in Organic Synthesis Francisco Sanchez-Madrid) J. McCusker / P. Coppens) (Barry Trost / Greg Fu / David Teller) • Innate Immunity and Toll Receptors • Organometallics in Catalysis (Alan Ezekowitz / Bruce Beutler / (Johannes de Vries / Daniel Mindiola) ______Jin Mo Park / Roland Strong) • Synthetic/Mechanistic Organometallic • Phagocytosis and Host Pathogen Chemistry Interactions (Peter Wolczanski / Paul Chirik / Vlad PHOTOSYNTHESIS ( / Sergio Grinstein / Grushin) BRYANT UNIVERSITY Steve Greenberg / Lee-Ann Allen / • Organometallic Lanthanide/Actinide SMITHFIELD, RI Rick Brown) Chemistry JUL 3-8, 2005 • Microbial Killing (Karsten Meyer) SABEEHA MERCHANT, CHAIR (William Nauseef / Anthony Segal / • Physical Methods R. DAVID BRITT, VICE CHAIR Elizabeth Ligeti / Tom DeCoursey) (Paul Pregosin) • Signal Transduction • Structure Function Analysis (Linda McPhail / Edgar Pick / Michael Yaffe / (A. William Rutherford / Kevin Redding / ______Phill Hawkins / Carl Nathan) Bridgette Barry / Junko Yano) • Emerging Technologies • Proteomics and Functional Genomics (John Curnutte / Alan Aderem / (Robert Blankenship / Norbert Rolland / ORIGINS OF SOLAR SYSTEMS Adrian Ozinsky) Dario Leister / Jeffrey Moseley / CONNECTICUT COLLEGE J. Thomas Beatty / Devaki Bhaya / NEW LONDON, CT ______Donald A. Bryant) JUN 26-JUL 1, 2005 • Proton and Electron Transfer JOSEPH NUTH, CHAIR (David Britt / Fevzi Daldal / Wolfgang Junge / LEE HARTMANN, VICE CHAIR PHOTOACOUSTIC & Frauke Baymann) PHOTOTHERMAL PHENOMENA • Protein Targeting and Biogenesis of • What Have We Learned from Extrasolar INTERNATIONAL CENTER FOR Complexes Planetary Systems? THEORETICAL PHYSICS (Eva-Mari Aro / Danny Schnell / (William Cochran / Artie Hatzes / Alan Boss) TRIESTE, ITALY Steven Theg / Maria Ghirardi / • The Genesis Mission: The Initial JUN 26-JUL 1, 2005 Richard Kuras / Himadri Pakrasi) Composition of the Solar System MLADEN FRANKO & JOSEF PELZL, CO-CHAIRS • CF Biogenesis and Regulation and ATP (Donald Burnett / Charles Hohenberg / 1 MASAHIDE TERAZIMA, VICE CHAIR Synthesis Robert Pepin) (David Kramer / Toru Hisabori / • Spitzer Observations of • Nanoscale Thermal Properties and Toshiharu Shikanai / Francis-André Wollman) Protoplanetary Disks Transport • Pigment and Cofactor Synthesis, Assembly (Michael Meyers / Dan Watson / Paola (Bernard Cretin / Azedine Hammiche / and Regulation D’Alessio) Ludwig Balk) (Elisabeth Gantt / Ayumi Tanaka / • Presolar Production of Short-Lived Radio • Nanoparticles Frédéric Barras / Stéphane Lobréaux / Isotopes (Jan Thoen / Egon Matijevic / B. Lounis / Niels-Ulrik Frigaard / Angela Falciatore) (Katrina Lodders / Roger Chevalier / Vladimir Zharov) • Antenna Protein Structure, Al Cameron) • Medical Applications Function and Regulation • Local Production of Short-Lived Radio (A.A. Oraevsky / Werner Faubel / (Richard Cogdell / Stefan Jansson / Isotopes Lihong V. Wang) Jean-David Rochaix / David Kehoe) (Guy Consolmagno / Eric Feigelson / • Linear and Nonlinear Laser Sound • Redox, Environmental Control and State Kevin McKeegan) (Vitaly Gussev / Sergey V. Egerev / Transitions • Constraints on Nebular Processes from Peter Hess / Otto Muskens) (Christine Foyer / Klaus Apel / Krishna Niyogi / Presolar Grains • Ultrafast and Transient Phenomena Aaron Kaplan / Giovanni Finazzi) (Sara Russell / Scott Messenger / Gary Huss) (David Hurley / M. Yamaguchi / O.B. Wright) • Structures of Photosynthetic Complexes • Disk Chemistry from Comets, Meteorites • Analytical Chemistry and Photochemistry (Marilyn Gunner / James Barber / and Spitzer (T. Autrey / Stephen Bialkowski / Nathan Nelson / Neil Hunter) (Geoff Blake / Paola Caselli / Bruce Fegley) Chieu. D. Tran / Silvia Braslavsky) • Disk Dynamics: The Formation and • Food Quality and Environmental Analysis Migration of Planets ______(Helion Vargas / Frans J.M. Harren / (Stuart Weidenschilling / Scott Kenyon / Dane Bicanic) Richard Nelson) • New Instrumental Developments • Comparative Planetology: (Mihai Chirtoc / Frank K. Tittel / Is the Earth Unique? J.G. Diebold / Ralph Meckenstock) (Geoffrey Taylor / Michael Drake) • Nondestructive Evaluation (Hassan Talaat / Andreas Mandelis / ______Gerd Busse) ______

visit the frontiers of science: www.grc.org PHYSICAL ORGANIC CHEMISTRY PLASMID & CHROMOSOME DYNAMICS • Bio Medical Applications HOLDERNESS SCHOOL (Haruma Kawaguchi / Yukio Nagasaki / PLYMOUTH, NH (see CHROMOSOME DYNAMICS) Eugenia Kumacheva / Guanghi Ma) JUN 26-JUL 1, 2005 • Self-Assembly GARY WEISMAN, CHAIR ______(Alex van Herk / Younan Xia / Eric Dufresne) R. STAN BROWN, VICE CHAIR ______• Stereochemistry; Industry & the Discipline POLYAMINES (Charles Perrin / Jay Siegel / CONNECTICUT COLLEGE Eusebio Juaristi / Ed Wasserman) NEW LONDON, CT POLYMERS (EAST) • Supramolecular Chemistry JUN 12-17, 2005 MOUNT HOLYOKE COLLEGE (Tyler McQuade / Cornelia Bohne / EUGENE GERNER & SOUTH HADLEY, MA Luis Echegoyen / Olaf Wiest / Lyle Isaacs / ANTHONY MICHAEL, CO-CHAIRS JUN 19-24, 2005 Silviu Balaban) LEENA ALHONEN & , CHAIR • Computational Chemistry MARGARET PHILLIPS, CO-VICE CHAIRS KAREN WOOLEY, VICE CHAIR (Richard Johnson / Weston Borden / David Giesen / Martin Saunders) • Physiological Roles • Functional Complex Macromolecules • Structure and Function I (Susan Gilmour / John Cleveland / (Tim Long / Takuzo Aida / Jean Frechet) (Roger Alder / Natia Frank / Darren Hamilton / Juahni Janne / Leonard Johnson) • Biological Routes to Polymers Lawrence Scott / Armin de Meijere / • Transport (Kristi Kiick / Richard Gross / David Tirrell) Rik Tykwinski) (Keiko Kashiwagi / Takeshi Uemura / • Polymeric Tissue and Drugs • Organometallic Chemistry Mattias Belting / Marie-Pierre Hasne / (Heather Maynard / Jeff Hubbell / (David Lemal / Guy Lloyd-Jones / Richard Poulin) Ruth Duncan) David am Ende / Hans Reich) • Mechanisms of Action: RNA / Translation • Molecular Brushes • Mechanisms, Intermediates, and Structure (John Atkins / Eric Westhof / Tairo Oshima / (William Brittain / Christine Ortiz / (Stephen Nelsen / Edward Clennan / Kazuei Igarashi) Sergei Sheiko) Steven Kass / William Leigh / • Mechanisms of Action: RNA Processing / • Reinventing Polyolefins Frances Cozens / Jonathan White) eIF-5A (Bruce Novak / Jim Stevens / Pete Fujita) • Enzymes (Albert Abbruzzese / Myung Park / • New Materials via Precise Polymer (Malcolm Forbes / John Richard / Sandro Valentini / Annette Kaiser) Synthesis Peter Petillo / Martin Tanner) • Mechanisms of Action: Translational (Tim Patten / Yves Gnanou / Jeff Pyun) • Structure and Function II Frameshifting / Antizyme • Bioinspired (Nancy Goroff / Joseph Lambert / (Senya Matsufuji / Phillip Farabaugh / (Helmut Ringsdorf / Sam Stupp / Virgil Percec) Thomas Bell / Anna Gudmundsdottir / Marvin Hackert / Philip Coffino) • Controlling Ionic and Kathleen Kilway / Peter Schreiner) • Mechanisms of Action: Signaling / Radical Polymerizations • Poster Session Talks Transcription (Stuart Rowan / Stan Penczek / (Stan Brown) (J.Y. Wang / Lisa Shantz / Ralf Baumeister / Mitsuo Sawamoto) Kirsi-Marja Oksman-Caldentey) • Polymers for Optoelectronics ______• Pathophysiological Functions: Cancer (Florian Schattenmann / Chris Ober / (Diane McClosky / David Feith / Ian McCulloch) Robert Casero / Frank Meyskens) • Nanoscale Organization of Polymers PLANT METABOLIC ENGINEERING • Pathophysiology / Clinical Applications (Doug Kiserow / Frank Bates / Ned Thomas) TILTON SCHOOL (Lo Persson / Carol Colton / Karen Doyle) • Novel Materials from Unconventional TILTON, NH • Debate: Therapeutic Strategies (Analogs Media JUL 10-15, 2005 Versus Target-Directed Drugs) (Bernadette Charleux / Joe DeSimone / ELEANORE WURTZEL, CHAIR (Carl Porter / Pro-Analogs: Patrick Woster / Markus Antonietti) ERICH GROTEWOLD, VICE CHAIR Heather Wallace / Laurence Marton / • Polymers at Surfaces Pro Target Directed: Ian Blagbrough / (Martin Moeller / Tom McCarthy / • Plant Chemistry and Ethnobotany Victor Levin) Tomek Kowalewski) (Jim Simon / Dennis Stevenson / • Macromolecular Nano-Objects Jorge Vivanco) ______(Valerie Shears / Axel Mueller / Craig Hawker) • Metabolic Profiling and Spectroscopic Tools ______(Richard Trethewey / Lloyd W. Sumner / POLYMER COLLOIDS Xuemin Wang / Ruth Stark) TILTON SCHOOL • Metabolic Flux Analysis and Modeling TILTON, NH POLYSACCHARIDES, CHEMISTRY OF (Andrew Hanson / David Gang / JUL 3-8, 2005 HONG KONG UNIVERSITY OF John Ohlrogge / Lukas Mueller) KOICHI TAKAMURA, CHAIR SCIENCE & TECHNOLOGY • Enzyme Structure and ALEX VAN HERK, VICE CHAIR HONG KONG, CHINA Assembly of Complexes JUN 5-10, 2005 (Deborah Delmer / Birger Møller / • Control in Particle Shape and SHIGENORI KUGA, CHAIR Brenda Winkel / Joseph Noel) Molecular Structure DEREK GRAY, VICE CHAIR • Metabolic Compartmentalization (Francoise Candau / Masayoshi Okubo / (Virginia Walbot / George Wagner / Bernadette Charleux) • Biosynthesis and Chemical Synthesis 1 Peter Facchini / Enrico Martinoia) • Challenge in New Industrial Processes and (Nick C. Carpita / Steven Ball / • From Genomes to Pathways Products Vincent Bulone) (Joe Chappell / Clint Chapple / (Mamoru Nomura / Ed Kostansek / • Biosynthesis and Chemical Synthesis 2 Kevin Walker / Yossi Hirschberg) Bedri Erdem / Dennis Larah) (Vincent Bulone / Shin-ichiro Nishimura / • Engineering the Senses • New Development in Nick C. Carpita / Jun-ichi Kadokawa) (Gad Galili / Eran Pichersky / Industrial Applications • Biodegradation and Francesca Quattrocchio) (Do Ik Lee / Wolf-Dieter Hergeth / Enzymatic Treatments 1 • Plants as Chemical Factories Charles McDonald) (Harry Brumer / Masahiro Samejima / (Dean Della Penna / Alisdair Fernie / • Novel Composite Colloids Antje Potthast) Jacqui Shanks / Craig Nessler) (Mitchell Winnik / Wen-Ying Chiu / • Biodegradation and • Emerging Technologies: Onder Pekcan / Jennifer Lewis) Enzymatic Treatments 2 A Window to the Future • Challenge in Process Engineering (Yoshiharu Nishiyama / Takuya Kitaoka / (Jay Keasling / Hal Alper / Bernhard Palsson) (Jose Asua / Mike Taylor / Andrew Klein) Harry Brumer) • Control in Dispersion Stability • Structure and Properties ______(Peter Lovell / Yilong Han / Darrell Velegol / (Antje Potthast / Yoshiharu Nishiyama / Theodorus van de Ven) John W. Brady / Stephen J. Eichhorn)

visit the frontiers of science: www.grc.org • Complex Formation • Protein Design • Cloud Observing Systems (Laurent Heux / Alain Buleon / (Donald Doyle / Homme Hellinga / (Susanne Crewell / Steven Platnick / Shinichi Kitamura / Bjorn T. Stokke) Brian Kuhlman / Lynne Regan / Steve Mayo / David Turner / David Donovan) • Colloids and Materials 1 Andreas Pluckthun) • Assimilation of Cloud Data (Bjorn T. Stokke / Masahisa Wada / • Natively Disordered Proteins (Arthur Hou / Tomislava Vukiæeviæ / Laurent Heux) (Trevor Creamer / Peter Wright / Keith Dunker) Angela Benedetti) • Colloids and Materials 2 • Membrane Proteins • Modelling and Observing Clouds and (Saad A. Khan / Francois Ravenelle / (Chris Hill / Robert Stroud / Radiation Dieter Klemm / Hiroyuki Yano) Harvey McMahon / Lukas Tamm / (Wojciech Grabowski / David Randall / • Colloids and Materials 3 Dinesh Yernool / Jim Bowie) Christian Jakob / Xiaoqing Wu) (Dieter Klemm / Lina Zhang / Saad A. Khan) • Proteins in Disease • Analyses of Cloud Feedbacks and (Gary Pielak / Ron Wetzel / Radiative Sensitivities ______Jonathan Weissman) (Hartmut Grassl / Alan Betts / Bruce Wielicki) • Protein Complexes and Networks (Volker Dotsch / John Kuryian / Lila Gierasch / ______Art Johnson / / Tania Baker) PROTEIN TRANSPORT • Closing Keynote Speaker ACROSS CELL MEMBRANES (Virginia Rath / Wendell Lim) COLBY-SAWYER COLLEGE RED CELLS NEW LONDON, NH TILTON SCHOOL JUN 12-17, 2005 ______TILTON, NH HARRIS BERNSTEIN & JUN 12-17, 2005 ROSS DALBEY, CO-CHAIRS LEONARD ZON, CHAIR JURGEN SOLL, VICE CHAIR QUANTUM CONTROL OF LIGHT AND MATTER JON MORROW, VICE CHAIR COLBY COLLEGE • Protein Targeting WATERVILLE, ME • Erythroid Gene Expression: Membrane (Lila Gierasch / Roland Beckmann / JUL 31-AUG 5, 2005 Proteins Ralph Henry / Olaf Schneewind) PAUL CORKUM, CHAIR (David Bodine / Peter Agre / Harvey Lodish / • Protein Import into Mitochondria PHILIP BUCKSBAUM & Edward Benz / David M. Bodine) (Trevor Lithgow / Toshiya Endo / DAVID TANNOR, CO-VICE CHAIRS • Erythroid Gene Expression: Globin / LCR Carla Koehler / Klaus Pfanner / (Douglas Engel / Douglas R. Higgs / Rosemary Stuart) • Opening Session Frank Grosveld / Gerd Blobel / • Translocation Across the (H. Rabitz) Emery Bresnik / Doug Engel / Peter Fraser) Bacterial Inner Membrane • Controllability • Membrane Rafts, Receptors and Dynamics (Tassos Economou / Arnold Driessen / (P. Brumer / R. Brockett / N. Khaneja / (Velia Fowler / Kai Simons / Kasturi Haldar / Matthias Muller / Tracy Palmer) C. Rangan / T. Weinacht) Athar Chisti / Mohandas Narla / • Protein Translocation Into and • The Role of De-Coherence Rainier Prohaska) Out of the ER (M. Scully / D. Lidar / E. Shapiro) • RBC Structure, Organization and (Reid Gilmore / Manu Hegde / Hidde Ploegh / • Attosecond and X-Rays Volume Control Yihong Ye) (S. Leone / F. Kaertner / M. Murnane / (Joseph Hoffman / Diana Gilligan / • Peroxisome Protein Import R. Kienberger) Clive Ellory / Phil Low / Ruby MacDonald / (Paul Lazarow / Ralf Erdmann / • Light Matter Interaction at its Extreme Joel Chassis / David Speicher / Yukio Fujiki / Steven Gould) (J. Kimble / L. Novotny) Stephen Goodman) • Transport to the Bacterial Cell • Control in Few Level Systems • Transcription Surface and Beyond (K. Bergmann / S. Rice) (James J. Bieker / Rick Young / (Hajime Tokuda / Peter Christie / • Wave Packets and Their Applications James J. Bieker / Nancy Speck / Vassilis Koronakis / Tony Pugsley / (T. Kobayoshi / R. Jones / D. Villeneuve) Patrick G. Gallagher / Joseph Prchal) Jan Tommassen) • Chemical Control • Red Cell Disorders • Protein Import into Chloroplasts (R. Levis / N. Schwentner / (David G. Nathan / David G. Nathan / (Colin Robinson / Ken Cline / Danny Schnell / V. Bonacic-Koutecky / G. Gerber / Y. Ohtsuki) Barry Paw / Gordon Stewart / Steve Theg) • Quantum Thermodynamics and Cooling Nancy Andrews / LuAnn Peters / • Insertion/Folding of Membranes Proteins (M. Raizen / R. Kosloff) Chang-Zheng Chen) (Gunnar von Heijne / Bill Dowhan / • Stem Cells Art Johnson / Andreas Kuhn / Bill Skach) ______(Stuart H. Orkin / Stuart H. Orkin / • “Non-Classical” Mechanisms of Protein Masayuki Yamamoto / Tariq Enver / Transport Across Cell Membranes Anna Rita Migliaccio / Margaret H. Baron) (David Andrews / Steve Dowdy / • Hematopoiesis RADIATION & CLIMATE Francoise Jacob-Dubuisson / Anna Rubartelli) (Roger K. Patient / Roger K. Patient / COLBY COLLEGE Stephan Karlsson / Kyunghee Choi / WATERVILLE, ME Bill Detrich / Toshio Suda / Elaine Dzierzak / ______JUL 24-29, 2005 Todd R. Evans) HOWARD BARKER & • Selected Talks from Poster Presentations ROBERT ELLINGSON, CO-CHAIRS (John Conboy / Gordon Grinder) PROTEINS WILLIAM COLLINS & HOLDERNESS SCHOOL PHILIP RUSSELL, CO-VICE CHAIRS PLYMOUTH, NH ______JUN 19-24, 2005 • Hydro-Radiative Climatology VIRGINIA RATH & FRANZ SCHMID, CO-CHAIRS (Dennis Hartmann / / CHRISTOPHER HILL & Syukuro Manabe) SECOND MESSENGERS & GARY PIELAK, CO-VICE CHAIRS • Absorption and Scattering PROTEIN PHOSPHORYLATION (Qiang Fu / Eli Mlawer / Klaus Pfeilsticker / KIMBALL UNION ACADEMY • Opening Keynote Speaker Micheal Mishchenko) MERIDEN, NH (Franz Schmid / Susan Lindquist) • Radiative Transfer I: Two-Stream Approxi- JUN 12-17, 2005 • and Dynamics I mation and Averaging JEFFREY BENOVIC & (Terry Oas / George Rose / Art Palmer / (James Coakley, Jr. / Warren Wiscombe & MARK VON ZASTROW, CO-CHAIRS / Andy Robertson / Philip Gabriel / Alexander Marshak) NATALIE AHN & Vijay Pande) • Radiative Transfer II: Dynamical Models BEVERLY ERREDE, CO-VICE CHAIRS • Protein Folding and Dynamics II (Thomas Ackerman / Petri Räisänen / (Charles Brooks / Dan Raleigh / Bernard Pinty / Dana Veron) • GPCR Signaling Rudi Glockshuber) • Radiative Transfer III: (Michel Bouvier / Thue Schwartz / Randy Hall) Innovative Remote Sensing • Regulation of GPCR / G Protein Signaling (Lee Harrison / Qilong Min / Anthony Davis) (Martin Lohse / Joann Trejo / Maurine Linder / Heidi Hamm / Lou Luttrell)

visit the frontiers of science: www.grc.org • Lipid Signaling STAPHYLOCOCCAL DISEASES • Experimental Design and Analysis for (Ken Harden / Pat Casey / Marc Lemmon) SALVE REGINA UNIVERSITY Accelerated Degradation Testing of • Heterotrimeric G Protein Regulation NEWPORT, RI Li-Ion Cells (Ken Blumer / Steve Lanier / AUG 21-26, 2005 (Ed Thomas / Brad Jones / David Siderovski / Michael Koelle / ARNOLD BAYER & JEAN LEE, CO-CHAIRS Gautham Parthasarathy) Henrik Dohlman) MATHIAS HERRMANN & • Chemometrics Applications to Systems • Structural Insight into Signaling HARALD LABISCHINSKI, CO-VICE CHAIRS Biology: Genomics, Proteomics, (John Sondek / John Tesmer / Brian Kobilka / Metabonomics and Lipomics Kris Palczewski / Steve Sprang) • Genomics and Proteomics under Relevant (Raymond Lam / Gregory Warnes / • Sensory Signaling Biologic Contexts David Deuwer) (Randy Reed / Hiroaki Matsunami / (Paul Dunman / Ken Bayles / Mark Smeltzer / ) Susanne Englemann) ______• Plenary Session • Bacterial Physiology and Metabolism (Mark von Zastrow / Jeffrey Benovic / Charles Linked to Pathogenicity Zuker / ) (Richard Proctor / Barry Bochner / STRESS PROTEINS IN GROWTH, • Signaling Integration / Networks Greg Somerville / Simon Foster) DEVELOPMENT & DISEASE (Natalie Ahn / Rama Ranganathan / • Regulation of Autolysis SALVE REGINA UNIVERSITY Ravi Iyengar / Alex Brown / Beverly Errede) (Markus Bischoff / Ambrose Cheung / NEWPORT, RI • Physiological Models of Signaling Kelly Rice / Christiane Goerke) JUL 17-22, 2005 (Marc Caron / Peter Mombaerts / • Staphylococcal Interactions with Host Cells DENNIS THIELE, CHAIR James Hurley / Tom Scanlan) (Michael Yeaman / Banhu Sinha / PETER WALTER, VICE CHAIR • Nuclear Sensors and Signaling Timothy Foster / Hattie Gresham) (Richard Fishel / David Toczyski / • Immune Responses to • Keynote Address Helle Ulrich / Craig Thompson) Staphylococcus Aureus (Elizabeth Craig) (Andreas Peschel / Thomas Hartung / • Proteotoxicity Gabriel Nunez / Arthur Tzianabos) ______(Bernd Bukau / Carol Gross) • New Concepts of Cell Wall • Stress in Cellular Compartments Structure and Function ( / Jonathan Weissman / (Henry Chambers / Brigitte Berger-Bachi / SMALL INTEGRIN-BINDING PROTEINS Jeffrey Brodsky) Maria Pinho / Brian Wilkinson) BIG SKY RESORT • Stress Signals in Transcription Activation • Community-Acquired Methicillin-Resistant BIG SKY, MT (Lea Sistonen / Akira Nakai / Staphylococcus Aureus (CA-MRSA) - SEP 11-16, 2005 Xinnian Dong / Michele Toledano) Molecular and Clinical Paradigms WILLIAM BUTLER & • Mechanisms of Stress Regulation (Barry Kreiswirth / Frank Lowy / LARRY FISHER, CO-CHAIRS (Richard Morimoto / John Lis / Robert Daum / Frank DeLeo) Caroline Jolly / James Goodrich) • Issues in Staphylococcus Epidermidis • Introduction to Integrins & • Stress Proteins and Survival (Gordon Archer / Michael Otto / the SIBLING Family (Ivor Benjamin / Jeffrey Robbins / Paul Fey / Wilma Ziebuhr) (William Butler / Dwayne Stupack / Kevin Morano / Doug Green) • Controversies/Hot Topics Tyra Wolfsberg) • Cell Biology of Stress (Benedicte Fourniere) • Shared Functions of SIBLINGs (Ron Kopito / Tso-Pang Yao) (Adele Boskey / Graeme Hunter / • Aging-Stress Axis Vincent Castronovo / Neal Fedarko / ______(Cynthia Kenyon / Lenny Guarente) Kalu Ogbureke) • Neurodegenerative Diseases and Stress • Osteopontin & Mineral Metabolism (Susan Lindquist / Liming Li) (Esben Sorensen / Cecilia Giachelli / STATISTICS IN CHEMISTRY & Masaki Noda / Keith Hruska) CHEMICAL ENGINEERING ______• Osteopontin in Cancer, Inflammation & MOUNT HOLYOKE COLLEGE Immunity SOUTH HADLEY, MA (Susan Rittling / David Denhardt / JUL 17-22, 2005 STRUCTURAL, FUNCTIONAL & Ann Chambers / Jeffrey Berman / BABATUNDE OGUNNAIKE, CHAIR EVOLUTIONARY GENOMICS Toshimitsu Uede / Harvey Cantor) DAVID HAALAND, VICE CHAIR BATES COLLEGE • Bone Sialoprotein & LEWISTON, ME Dentin Matrix Protein 1 • Statistical Challenges in Systems Biology JUN 19-24, 2005 (Marc McKee / Jaro Sodek / (Age Smilde / Steve Brown / Sijmen de Jong) KEVIN WHITE, CHAIR Jane Aubin / Anne George) • Exploration and Resolution Methods EUGENE KOONIN, VICE CHAIR • The CCN Family Applied to Analysis of Protein Processes (Stephen Lam / Lester Lau / (Anna de Juan / Marcel Maeder) • Functional Genomics I: Karen Lyons / David Brigstock) • Analysis of Multivariate Images with Developmental Genomics • DMP1 & Dentin Sialophosphoprotein Applications to Process Monitoring and (Eric Siggia / Sean Eddy / Alan Michelson) (Lynda Bonewald / Jian Feng / Control • Proteins and Proteomes I: Mary MacDougall / Chunlin Qin) (John MacGregor / Mike Piovoso / Jay Lee) Predicting Protein Interactions • DSPP, Matrix Extracellular • Multi-Way Analysis of Metabonomics and () Phosphoglycoprotein (MEPE) & Hot Topics NMR Data: Challenges and New Results • Functional Genomics II: Approaches to (Michel Goldberg / Taduru Sreenath / (Rasmus Bro / Willem Windig / High Throughput Genetic Screening and Peter Rowe / Thomas Brown) Johan Westerhuis) Phenotyping • Banquet Talk - Snake Venoms: Integrins, • Applications of Statistics to Multiscale (Kevin White) Disintegrins & MMPs Systems • Evolution of Genome Complexity (Larry Fisher / Jay Fox) (Richard Braatz / Bhavik Bakshi / (Eugene Koonin / Chris Adami / Derrick Rollins) Richard Lenski / Michael Lynch) • PharmID: Pharmacophore Identification ______• Proteins and Proteomes II: Structural using Gibbs Sampling Genomics and Novel Protein Functions (Stan Young / Aaron Owens) (Doug Brutlag / Alexei Murzin / • Comprehensive Batch Process Models Janet Thorton / Chris Lima) with In-Situ Spectroscopic Measurements • Proteins and Proteomes III: and Calorimetry Macromolecular Complexes (Paul Gemperline / Sara Rutan / Dora Kourti) (Peer Bork) • Comparative Genomics (Yoav Gilad / Lawrence Hurst / Alex Kondrashov / Hunter Fraser)

visit the frontiers of science: www.grc.org • Functional Genomics III: THREE DIMENSIONAL TOXICOGENOMICS Chemical Genomics ELECTRON MICROSCOPY COLBY-SAWYER COLLEGE (Joel Bader) COLBY-SAWYER COLLEGE NEW LONDON, NH • Comparative Functional Genomics NEW LONDON, NH JUN 5-10, 2005 (Inna Dubchak / Arend Sidow / JUN 12-17, 2005 DANIEL LIEBLER & Adam Arkin / Dan Rokhsar) KENNETH DOWNING, CHAIR LEONA SAMSON, CO-CHAIRS & CINDY AFSHARI, VICE CHAIR ______PHOEBE STEWART, CO-VICE CHAIRS • Keynote Session • Opening Session: “Whither (Molecular) (Leona Samson / Daniel Liebler / Electron Microscopy?” Chris Bradfield) SUPRAMOLECULES & ASSEMBLIES, (Kenneth Downing / Ray Stevens / • Serum Proteomics and Markers of Toxicity CHEMISTRY OF Keiichi Namba) and Disease COLBY COLLEGE • High Resolution of Structures with (Tim Veenstra / Daniel Chan / Robbert Slebos) WATERVILLE, ME High Symmetry • Toxicogenomics and Chromatin Structure JUN 12-17, 2005 (Tim Baker / Phoebe Stewart) (Bernard Futscher) DAVID THOMPSON, CHAIR • Better Resolution of Structures with • New Technologies to Assess Protein DIRK KURTH & Low Symmetry Modifications and Toxicity JOHN TEXTER, CO-VICE CHAIRS (Melissa Jurica) (Serrine Lau / Forrest White / Natalie Ahn / • Molecular and Cellular Tomography Mark Bedford / Dennis Petersen) • Self-Assembly Using Viral Templates (Manfred Auer) • Genetic Susceptibility to Disease (Jack Johnson / Chad Mirkin) • Poster Presentations and Discussion I (Cynthia Afshari) • Nanostructure Design and Reactivity Using (Teresa Ruiz) • Metabonomics to Understand Chemical Nucleic Acid-Based Templates • New Software Tools Toxicity (Matt Kanan / Chengde Mao / Andy Ellington) (Jose-Maria Carazo) (Craig Thomas / James Willet / • Dynamic Supramolecular Systems • Instrumentation and John Lindon / Don Robertson) (David Reinhoudt / Jeremy K.M. Sanders / Specimen Preparation Advances • Genomic Approaches to Predictive Rint Sijbesma) (Bridget Carragher) Toxicology • Molecular Recognition & Interfacial • Applications in Building and Refining (Ray Tennant / James Stevens / Alison Vickers Phenomena in Crystallization Hybrid Structures / Lisiane Meira) (Bart Kahr / Alain Brisson / Robert Tampe) (Chris Akey) • Late-Breaking Research from • Hybrid Structures Emerging from Novel • Poster Presentations and Discussion II Submitted Abstracts Amphiphilic Materials and (Masahide Kikkawa) (Ben Van Houten / Cheryl Walker) Biomacromolecules • Functional Genomics in Model Systems (Charles Mioskowski / Wolfgang Meier / (Marc Vidal) Matt Tirrell) ______• The “Next Generation” (Christine Keating / Sarah Keller / ______Tyler McQuade / Sergei Sheiko / TISSUE REPAIR & REGENERATION Francesco Stellacci / Andrew Taton / COLBY-SAWYER COLLEGE Marcus Weck) NEW LONDON, NH TUBERCULOSIS DRUG DEVELOPMENT • Structure & Dynamics of Phase Separated JUN 19-24, 2005 KIMBALL UNION ACADEMY Lipid Domains in Biology JUDITH ABRAHAM & MERIDEN, NH (Paula Booth / Nancy Thompson / PAUL MARTIN, CO-CHAIRS JUL 3-8, 2005 Watt Webb) JACK GAULDIE, VICE CHAIR JAMES SACCHETTINI, CHAIR • Self-Assembly and Controlled Disassembly VALERIE MIZRAHI, VICE CHAIR in Drug and Gene Delivery • Genetically Tractable Models of Repair (Frank Szoka / Henry Kopecek) (Paul Martin / Mike Galko / Mike Redd / • Opening Session • Biological Applications of Nanopaticles, Howard Chang) (James Sacchettini / Barry Furr) Dendrimers & Nanogels • Cell Migration • Persistence Factors as Drug Targets (Sönke Svenson) (Pierre Coulombe / James Nelson / (Eric Rubin / David Russell / Bill Parks / Ilene Gipson) John Blanchard / Laura Via) ______• Angiogenesis • Drugs Targets and Resistance (Catherine Nobes / Kairbaan Hodivala-Dilke / (Valerie Mizrahi / William Jacobs) Ann Daugherty) • High Throughput and Virtual Screening • Inflammation (Ken Duncan / Tanjore Balganesh / THIN FILM & CRYSTAL GROWTH MECHANISMS (Joe Leibovich / William Muller / Thomas Keller / Jose Garcia-Bustos) MOUNT HOLYOKE COLLEGE Luisa DiPietro) • Targeting Dormant Bacteria SOUTH HADLEY, MA • Regeneration (Douglas Young / John McKinney) JUN 26-JUL 1, 2005 (Ken Muneoka / Elly Tanaka / • Structure-Based Approaches MELISSA HINES, CHAIR J.C. Izpisua-Belmonte) (Kurt Krause / Tom Albers / PETER VEKILOV, VICE CHAIR • Neurorepair Satheesh Palaninathan) (Anne Logan / Stephen Davies / • Lead Optimization • Quantum Dots and Semiconductor Growth Patrick Anderson / Larry Benowitz) (Melvin Spigelman / Kim Lewis / (Oliver Schmidt / Vivek Shenoy / • Growth Factors and Hormones in Repair Clifton Barry / Koen Andries) Jeff Drucker) (Judy Abraham / Sabine Werner / • Animal Models • Biological Control of Crystallization Gillian Ashcroft / Jeff Hubbell) (William Bishai / Joanne Flynn / Ann Lenaerts) (Naomi Chayen / Peter L. Davies) • Fibrosis • Clinical Trials and • Thin Film Growth Dynamics (Jack Gauldie / Mark Ferguson / New Therapeutic Strategies (Marcel J. Rost / Kristin Fichthorn / Snorri Thorgeirsson) (Denis Mitchison) Jonah Erlebacher) • Stem Cells; Clinical Applications • Dynamics of Crystal Growth (Paul Martin / Richard Bucala / (Michael D. Ward / Miquel Salmeron) ______Yann Barrandon / Mike Longaker) • Biomineralization (Daniel E. Morse) • Control of Nucleation ______(Bruce A. Garetz) • Organic Crystals (Michael Doherty) • Hot Topics and Contributed Talks (Melissa A. Hines)

visit the frontiers of science: www.grc.org • VISUALIZATION IN SCIENCE & EDUCATION B12 in Medicine and Toxicology I ZEOLITIC & LAYERED MATERIALS THE QUEEN’S COLLEGE (Margareta Törnqvist / William Watson / MOUNT HOLYOKE COLLEGE OXFORD, UK David Smith / Sergey Fedosov) SOUTH HADLEY, MA JUL 3-8, 2005 • Protein JUL 3-8, 2005 GEORGE LISENSKY & (Christoph Kratky / Catherine Drennan) KENNETH BALKUS, CHAIR • PETER MAHAFFY, CO-CHAIRS B12 and B12-Like Mechanisms ROBERT BEDARD, VICE CHAIR ROY TASKER & (Martin Newcomb / Neil Marsh) • • CHRISTOPHER WATTERS, CO-VICE CHAIRS B12 in Medicine and Toxicology II Synthesis (R. Carmel / Robert Kadner / Douglas Collins) (Stacey Zones / Greg Lewis / • Seeing and Understanding: An Overview • Cobalamin Russel Morris / Sandeep Dingrah) (Mary Shultz / Michael King / Neil Stillings) (Gabi Diekert / Christoph Holliger / • Mesoporous Materials • Haptics and Visualization at the Rowena Matthews / Rolf Thauer) (Serge Kaliaguine / Victor Lin / Kai Landskron) Micro- and Nano-Scale • Corphins • Membranes (Barbara Tversky / Gail Jones / (Bernhard Jaun) (Yushan Yan / Tina Nenoff / John Falconer) Tim Herman / Miriam Reiner) • Model Systems • Catalysis • Better Ways of Seeing and (Chemical and Computational) (Ahmad Moini / Peter Smirniotis / CY Chen) Understanding Science (Leo Radom / Rudi van Eldik / • Adsorption/Diffusion (Peter Atkins / George Whitesides / Kurt Warnke / Yoshio Hisaeda) (Scott Auerbach / Randall Snurr / Shaaron Ainsworth) • Perspective and Prospects Robert Thompson) • Visualizing Biological Complexity (Bernhard Kräutler / • Unusual Frameworks (Chris Watters / Benno Schwikowski / Jorge Escalante-Semerena / Ruma Banerjee) (Paul Wright / Omar Yahgi / Steve Suib) Malcolm Campbell / Kathy Takayama) • Poster Highlights • Visualization at the Micro-and Nano-Scale: ______(Robert Bedard) Research and Education (Zafra Lerman / Jillian Buriak / Patti Schank) ______• Seeing and Understanding with New Tools X-RAY PHYSICS and Environments COLBY-SAWYER COLLEGE (Pat Hanrahan / Niescja Turner / NEW LONDON, NH Ruth Chabay) AUG 7-12, 2005 • Best Practices in Visualization: ROBERT FEIDENHANSL, CHAIR Pedagogical and Cognitive Perspectives KENNETH FINKELSTEIN, VICE CHAIR (Loretta Jones / Roy Tasker / John Moore / John Gilbert) • Time Resolved Experiments • Seeing and Understanding our World and (Michael Wulff / Josef Feldhaus) Universe • New Sources (Henny Kramers-Pals / David Uttal / (Jochen Schneider / Ronald Ruth / Rosalind Grymes) David Moncton / Phillipe Zeitoun) • Resonant Scattering/Magnetism ______(Gerrit van der Laan / Peter Abbamonte) • Ultra Fast Science (Adrian Cavalieri / Kelly Gaffney /

VITAMIN B12 & CORPHINS Antoine Rousse / Lin Chen) THE QUEEN’S COLLEGE • Nano Science OXFORD, UK (Tetsuya Ishikawa / Harun Solak) SEP 18-23, 2005 • Coherent Applications BERNARD GOLDING, CHAIR (Gerhard Grübel / Stefan Eisebitt / WILFRED VAN DER DONK, VICE CHAIR Steve Wilkins / Lorentz Stadler) • Life Science • Cobalamin in Context (Alberto Bravin) (Ebba Nexø / Wolfgang Buckel / • Applications Tetsuo Toraya) (Harald Sinn / Marco Di Michiel)

75 years at the frontiers of science

In 2006, the Gordon Research Conferences will be celebrating its 75th Anniversary.

In the coming year, GRC will be developing a commemorative publication and web site chronicling our long and exciting history at the frontiers of science. Come to a GRC and see what all the excitement’s about. Be a part of history!

If you are interested in helping to preserve GRC’s history, please contact our director, Dr. Nancy Ryan Gray (401-783-4011), about sponsorship opportunities.

visit the frontiers of science: www.grc.org POSITIONS OPEN POSITIONS OPEN FACULTY POSITIONS in bioinformatics, medical image analysis, and visualization. The Department of Biomedical Informatics (BMI) Classified Advertising of the Ohio State University College of Medicine and Public Health (website: http://bmi.osu.edu/) The University of Connecticut School of Dental seeks applications for tenure-track faculty positions at Medicine is seeking an outstanding academic leader all levels. to direct its newly formed research Center for We seek broadly trained scientists with computa- Thomas Edison Regenerative Medicine and skeletal Development. tional research programs of biomedical relevance and 1847–1931 The DIRECTOR is expected to develop a Center collaborative potential. Scientists with research com- Founder of Science that will be recognized as a worldwide leader in plementing current efforts are of particular interest. research in the rehabilitation and regeneration of Interests within BMI include: biomedical image craniofacial tissue from basic, translational to clinical processing and quantification, comparative genomics, research. The focus of the established faculty in the phylogenetics, pharmacogenomics, cancer, protein Center spans the molecular regulation of skeletal and structure, promoter and chromatin analysis, and high craniofacial development, the creation of biomate- performance and grid computing. BMI has strong rials suitable for craniofacial and skeletal repair and collaborations in radiology, pathology, cardiology, regeneration, and the translational and clinical use of cancer, and automatic microscopic imaging. BMI emerging approaches to restorative, prosthetic, and offer opportunities to work in a rich multidisciplinary reconstructive dentistry. environment. For full details on advertising rates, deadlines, mechanical The person selected for this position will interact Applicants must have a proven record of publica- requirements, and editorial calendar, go to with an established, highly productive, and multi- tions in leading peer-reviewed journals and demon- www.sciencecareers.org and click on How to Advertise, disciplinary skeletal and craniofacial biology program strated potential to obtain extramural funding. or call one of our representatives. at the University of Connecticut Health Center Senior-level applicants (ASSOCIATE PROFES- (UCHC), consisting of 15 investigators in the SOR and above) should have extramural funding. United States & Canada School of Medicine and 11 in the School of Dental Teaching duties will include a course in an area of Medicine. A generous startup package will be specialty. E-mail: [email protected] available as well as considerable contemporary Applicants should send curriculum vitae, brief Fax: 202-289-6742 research facilities. Two additional vacant positions statements of research and teaching interests, copies jill downing also are part of the program to build a critical mass of representative publications, and have four letters of researchers in tissue repair and regeneration. of reference sent to: Search Committee, Bio- (CT, DE, DC, FL, GA, MD, ME, MA, NH, NJ, NY, NC, The ideal candidate should be a strong leader with medical Informatics, The Ohio State University, PA, RI, SC, VT, VA) research emphasis in any of the following areas: 3184 Graves Hall, 333 W. 10th Avenue, Colum- Phone: 631-580-2445 craniofacial biology, the design of scaffolds for tissue bus, OH 43210. kristine von zedlitz engineering, craniofacial translational research, or Recruitment is open immediately and will be the testing of experimental procedures in animals or ongoing until positions are filled. (AK, AR, CA, CO, HI, ID, IL, IA, KS, LA, MN, MO, MT, patients, along with a solid record in grant support NE, NV, NM, ND, OK, OR, SD, TX, UT, WA, WI, WY) and publications and extensive mentoring skills. DEPARTMENT CHAIR Phone: 415-956-2531 Interested candidates should send a letter of Molecular, Cellular, and interest and curriculum vitae to: Developmental Biology beth dwyer University of Colorado at Boulder (AL, IN, KY, MI, MS, OH, TN, WV and Internet Sales) Dr. S. Reisine, Chair of the School of Dental Medicine We seek an internationally recognized scientist Phone: 202-326-6534 University of Connecticut with a highly regarded research program and a emnet tesfaye 263 Farmington Avenue, MC3910 commitment to graduate and undergraduate educa- Farmington, CT 06030 tion. Molecular, Cellular, and Developmental Biol- (Line Advertising) ogy (MCDB), website: http://mcdb.colorado. UCHC is an Equal Opportunity Employer, Minorities/ Phone: 202-326-6740 edu/, is a distinguished and diverse department Females/Veterans/Persons with Disabilities. daryl anderson with a current faculty of 30. Attractions of this FACULTY POSITIONS: position include the expectation that during the next (Canada and Meetings and Announcements) few years, substantial new departmental resources Phone: 202-326-6543 ASSISTANT PROFESSOR (Tenure Track/Ph.D. Required) will flow from a drug developed using the SELEX LECTURER process invented in MCDB and recently approved by Europe & International (Nontenure Track/ the FDA. In addition the University has embarked Master_s Degree Required) on a parallel Biotech Initiative that will bring several E-mail: [email protected] Coastal Carolina University new scientists to the Boulder community. The new Fax: +44 (0) 1223-326-532 _ chair will have an unusual opportunity to implement The successful candidates primary responsibility a vision of the future for MCD Biology in an tracy holmes will be teaching/coordinating an interdisciplinary outstanding academic environment. science course for nonmajors, but teaching assign- Phone: +44 (0) 1223-326-525 _ Appointment could begin as early as fall 2005. ments in the individuals disciplines are possible. We Please send curriculum vitae, description of research gareth stapp are especially interested in applicants with a demon- plans, and the names of three references by mail to: Phone: +44 (0) 1223-326-527 strated interest in innovative methods of science Jane Richards, University of Colorado, 347 education and who will collaborate with faculty from helen moroney UCB, Boulder, CO 80309-0347, U.S.A., or as the Sciences and other Colleges within the Univer- PDF files by e-mail: [email protected]. Phone: +44 (0) 1223-326-528 sity to further define and develop the direction of If sending by mail, please include PDF files on disk this program. While the position will be housed in christina harrison in addition to printed originals. Dossiers will be the Department of Chemistry and Physics, applicants considered upon receipt until position is filled. Phone: +44 (0) 1223-326-510 with a background in any natural science will be The University of Colorado at Boulder is committed to jason hannaford considered. diversity and equality in education and employment. Applicants should submit a letter of interest, Phone: +81 (0) 52-777-9777 curriculum vitae, and contact information for three RESEARCH ASSISTANT PROFESSOR. The references to: Louis E. Keiner, University Hall Department of Physical Medicine and Rehabilita- To subscribe to Science: 207, Department of Chemistry and Physics, tion, University of Kentucky College of Medicine is In U.S./Canada call 202-326-6417 or 1-800-731-4939 In the rest of the world call +44 (0) 1223-326-515 Coastal Carolina University, P.O. Box 261954, seeking to fill a Research Assistant Professor position Conway, SC 29528-6054. Review of candidates in the area of spinal cord injury. Successful candi- will begin on March 1, 2005, and continue until the dates are expected to develop and maintain innova- Science makes every effort to screen its ads for offensive positions are filled. tive, nationally recognized research programs. and/or discriminatory language in accordance with U.S. and Coastal Carolina University is a growing, state- Interested applicants should submit curriculum non-U.S. law. Since we are an international journal, you may supported liberal arts institution where the emphasis vitae, statement of research interests, and contact see ads from non-U.S. countries that request applications from is on undergraduate education, and growing impor- information for three references to: Joe E. Springer, specific demographic groups. Since U.S. law does not apply to other countries we try to accommodate recruiting practices of tance is placed on faculty mentored student research Ph.D., Physical Medicine and Rehabilitation, other countries. However, we encourage our readers to alert projects and public services. Coastal Carolina Uni- University of Kentucky College of Medicine, us to any ads that they feel are discriminatory or offensive. versity is located approximately nine miles from Lexington, KY 40536-0284; e-mail: jspring@ Myrtle Beach, South Carolina, and enrolls more uky.edu. Female and minority candidates are encouraged to than 7,000 students. Coastal Carolina University is an apply. The University of Kentucky is an Equal Opportunity Equal Opportunity/Affirmative Action Employer. University. 770 4 FEBRUARY 2005 VOL 307 SCIENCE www.sciencecareers.org

CHAIR Department of Pharmacology Case Western Reserve University School of Medicine Nominations or applications are invited from established, dynamic scientists with a creative vision for the position of Chair of the Department of Pharmacology at the Case Western Reserve University School of Medicine. The Department has great tradition, and it has interactive faculty, outstanding graduate programs, facilities, space and scientific and educational activities. With a new leadership in the School of Medicine, the Department has been targeted for signifi cant growth to become one of the leading departments of Pharmacology in the country. The successful candidate should have an outstand- ing record of scholarly achievement and a record of leadership, mentoring and administrative abili- ties and a commitment to expand and enhance the Department to a nationally prominent role. Applicants should submit a letter and a statement of their research, teaching, service and admin- istrative experience, as well as their previous mentoring experience; their legacy in their own institution in building interdisciplinary programs and resources; a C.V. and a list of publications. Nominations and/or applications should be e-mailed to [email protected]. For additional information, visit http: //pharmacology.cwru.edu/. For questions or additional information you may call Toni Scarpa at 216-368-5298. Case Western Reserve University is an Equal Opportunity/Affi rmative Action Employer.

www.ars.usda.gov Interdisciplinary: Supvy Research Plant Pathologist; Supvy Microbiologist; Supvy Entomologist; GS-434/403/414-14/15 Salary Range: $85,123.00 - $130,173.00 PA

The Crop Diseases, Pests and Genetics Unit at the San Joaquin Valley Agricultural Sci- ences Center in Parlier, California is seeking a permanent, full-time Research Leader to: lead a multi-disciplinary team of scientists to determine the epidemiology of and develop strategies to manage, diseases caused by inva- sive plant pests (including plant pathogens and insects). The research will initially be focused on Xylella fastidiosa (Xf) strains and its insect vectors, especially the glassy-winged sharpshooter (GWSS).

For more details and application directions, see www.afm.ars.usda.gov/divisions/hrd/ index.html Announcement number is ARS- X5W-0152. Announcement closes March 11, 2005. For questions you may contact Dr. Ed Civerolo on 559-596-2702 or e-mail: [email protected]. U.S. Citizen- ship is required.

USDA, ARS is an Equal Opportunity Employer and Provider. Three Research Group Leaders in Stem Cell Biology

A new Stem Cell Research Initiative is planned for the Institute of Reproductive and Developmental Biology. Three tenure - track positions are available for outstanding scientists to develop independent research programmes in fundamental aspects of stem cell biology.

We are particularly interested in enthusiastic young scientists working on embryonic stem cells but other areas of stem cell research will be considered.

There will be opportunities to study human embryology and embryonic stem cells in conjunction with the In Vitro Fertility Clinic at the Hammersmith Hospital. The IRDB also has close links with the MRC Clinical Sciences Centre providing excellent core facilities and opportunities for collaborative research with the Epigenetics and Development cluster of groups. Honorary appointments in the CSC may be available if appropriate.

The IRDB currently comprises 15 interdisciplinary research groups focusing on cellular and molecular aspects of reproductive science. The Institute is located in a new modern building with excellent well-equipped laboratories. Information about the IRDB and current research programmes can be found in www1.imperial.ac.uk/medicine/about/ institutes/irdb Additional core services, including transgenic and embryonic stem cell support, microarray and bioinformatic facilities and advanced imaging are available elsewhere on the Hammersmith campus.

Group leaders will be appointed as University Lecturers with a salary in the range £33,817 - £37,772 per annum or up to £42,224 for exceptional candidates. A generous start-up package to establish an independent group will be available, especially to individuals with existing Fellowship support. The positions will be for three years in the first instance but are expected to lead to permanent academic appointments, subject to satisfactory productivity.

Informal enquiries may be made to Malcolm Parker, Director of Research, IRDB, , Du Cane Road, London W12 0NN. Email [email protected]

For an application pack please contact the Human Resources Division, Imperial Collge London, Hammersmith Campus, Commonwealth Building, Du Cane Road, London W12 ONN quoting reference number HJ796.

Alternatively an application form and job description may be obtained from the following web link http://www.imperial.ac.uk/employment/index.htm and sent to the above address.

Applications should include a 2 - 4 page description of current and future interests, a CV and the names of three referees. There is no formal deadline for applications although we are seeking to fill the positions in 2005.

Faculty Position in Breast Cancer Biology Department of Physiology - Division of Pathology The Department of Physiology invites applications for a full-time tenure-track appointment at the Assistant/Associate Professor level in a contemporary area of breast cancer biology. Areas of interest include, but are not limited to, cell or animal models of tumor transformation, progression, and metastasis; cancer genetics; breast cancer pathogenesis and molecular pathology; normal Located in Westminster, CO, Myogen is engaged in the and cancer stem cell biology; and the role of heterotypic cell interactions in discovery, development and commercialization of small the normal and cancerous breast. Candidates having expertise with genomics molecule therapeutics for the treatment of cardiovascular and/or proteomics, the genetic manipulation of animal models, as well as disorders. We currently have excellent opportunities for molecular, live cell, or whole animal imaging are especially encouraged to several highly motivated professionals to join our team in apply. It is anticipated that the candidate will contribute to collaborative the following key roles: research efforts in the area of breast cancer biology. Opportunities exist to participate in a NIEHS/NCI sponsored Research Center on Breast Cancer and the Environment. Candidates must hold a Ph.D., M.D., or equivalent RESEARCH ASSOCIATE doctoral or professional degree, have postdoctoral experience, and be In Vivo - Pharmacology able to demonstrate the potential to develop a vigorous externally funded CIENTIST research program. The candidate is expected to contribute to graduate and/or S professional teaching in an area of human or animal pathology. Cardiac Biology Applicants should submit as a single pdf file, a complete c. v. and SCIENTIST statement of current/future research plans. Applicants should also request Toxicology letters of recommendation from three individuals who can evaluate their accomplishments and future potential for research and teaching. Applications SCIENTIST should be electronically submitted March 5, 2005 to: In Vitro - Pharmacology Sandra Haslam, Ph.D. Chair, Breast Cancer Search Committee Myogen provides a competitive Department of Physiology – Division of Pathology compensation and benefits package. Michigan State University East Lansing, MI 48824-3320 View complete job descriptions and apply online at: [email protected]

Michigan State University is an Equal Opportunity/Affi rmative Action Equal Opportunity Employer Employer. Handicappers have the right to request and receive reasonable accommodation. DEPUTY MANAGER, EDUCATION AND SCIENCE & TECHNOLOGY, SUSTAINABLE DEVELOPMENT DEPARTMENT

The IDB is moving to The Inter-American Development Bank, the largest and leading source of financing for regional development in Latin America and the strengthen significantly Caribbean, based in Washington D.C., invites applications for the new position of Deputy Manager for Education and Science & Technology, its operations aimed in the Sustainable Development Department. at supporting the The Deputy Manager for Education and Science & Technology is a senior member of the management team of the Sustainable Development Department with development of national systems main responsibility for providing leadership and effective management in the areas of education, science & technology, including information and communication of innovation in the region. technology from the vantage point of an Institution with the mandate and resources to develop relevant projects and initiatives. In doing this, it draws from a long The Deputy Manager for Education and Science To qualify, the applicant should have: tradition of support for research, graduate education and & Technology will: • A Ph.D. in science, engineering or social sciences. technological development through • Oversee and coordinate technical support for operations in the areas of science and technology • Familiarity with and a record of achievement in issues of scientific and technological development, as linked with education, the projects that have benefited development, information and communication technologies for development and education. In accomplishing these tasks, the Deputy Manager will focus on working in close coordination with economics of innovation, competitiveness and productivity growth. numerous government institutions, the Departments of the Bank directly responsible for lending and regional cooperation aimed at • Proven abilities as a leader and manager of a creative and highly business and universities across maximizing development effectiveness in IDB operations. trained team. Latin America and the Caribbean. • Provide leadership within the organization to enhance and mainstream innovative policies as a key • Demonstrated ability to provide intellectual leadership and strategic component of national development agendas and IDB financial and technical assistance activities. direction. • Strong interpersonal and communication skills. In today’s global, knowledge based • Be a focal point for the development of strategies, policies, operational guidelines and applied research agendas of the IDB and play a leading role in their dissemination and implementation both • At least 15 years of experience (including significant international economy, science and technology within the Bank and in the context of larger policy dialogue sustained with governments, the private experience) in positions of increasing responsibility in the have become, more than ever sector, civil society and academia. management of government, business, academic or development before, critical to economic growth, • Interface with IDB’s partners, clients and stakeholders to promote coherence and effectiveness in organizations. competitiveness and quality of life. international assistance to scientific, technological and educational development as well as to • Proficiency in English and Spanish required. Working knowledge of project the Bank’s leadership in these areas. Portuguese and/or French is desirable.

To ensure strong economic If you are interested and qualify, please send your resume by February 28, 2005 growth and reduction of poverty Preferably by e-mail to: [email protected] in the future, the region must Or by mail to: significantly strengthen its Manager, Human Resource Department performance with respect to all IDB, 1300 New York Avenue, NW , Stop E401 • Washington, DC 20577 aspects of the innovation system. The IDB offers internationally competititive benefits and compensation, including relocation. You must be a citizen of one of the IDB Member Countries in order to qualify for any type of employment at the IDB. IDB Member Countries

• Argentina • Austria • Bahamas • Barbados • Belgium • Belize • Bolivia • Brazil • Canada • Chile • Colombia • Costa Rica • Croatia • Denmark • Dominican Republic • Ecuador • El Salvador • Finland • France • Germany • Guatemala • Guyana • Haiti • Honduras • Israel • Italy • Jamaica • Japan • Mexico • Netherlands • Nicaragua • • Panama • Paraguay • Peru • • Slovenia • Spain • Suriname • Sweden Inter-American Development Bank • Switzerland • Trinidad & Tobago • United Kingdom • United States • Uruguay • Venezuela •

Faculty Position Marine or Environmental Microbiology University of California, San Diego Department of Molecular Biology and Scripps PALEOBIOLOGY Institution of Oceanography The Department of Geological and Environmental Sciences at Stanford http://biology.ucsd.edu/ University invites applications for a tenure-track faculty appointment at http://www-sio.ucsd.edu/ the Assistant Professor level in the area of Paleobiology. We are looking for a person with a demonstrated research record and who is committed The Scripps Institution of Oceanography and the Section of Molecular to quality undergraduate and graduate teaching. While we will consider Biology in the Division of Biological Sciences, University of California, applications from individuals in all areas of paleobiology, emphasis will San Diego, invite applications for an Assistant (tenure-track), Associate be placed on candidates who have an understanding of the broad evolution (tenured), or Full Professor (tenured) in the area of marine or environmen- of life on Earth and research experience in one or more of the following tal microbiology. The successful candidate will hold joint appointments areas: (1) the origin and early evolution of life, (2) relationships between in the two departments as part of a developing program in microbiology. the evolution of life and the major physical processes and events in Minorities and women are encouraged to apply. Earth history, (3) external driving forces of major biological extinctions and radiations, and (4) climatic-ocean history and the evolution of the The successful candidate will use molecular approaches to the study of ancient marine biota. Interdisciplinary approaches are of special interest, fundamental problems in prokaryotic organisms, preferably from a marine but applicants must have a clear grounding in geology. We are seeking an environment. Areas of interest include but are not limited to: Archaeal individual who applies fundamental biological and geological principles, biology, microbial interactions in biofi lms, symbioses, or pathogenesis, quantitative data, and fi eld-based studies to characterize and model and genomic approaches to microbial evolution and diversity. The suc- biological evolution. cessful candidate is expected to develop or continue a vigorous research program of relevance to the goals of both departments and to participate in Although the position will remain open until fi lled, applications, including undergraduate and graduate teaching in microbiology. Salary will be com- a curriculum vitae, a statement outlining research and teaching interests mensurate with level of appointment and based on the UC salary scale. that would materially contribute to related programs in the School of Earth Sciences, and the names and addresses of three referees, should Complete applications received by March 15, 2005 will be assured of be sent by April 1, 2005, to: Paleobiology Search Committee, Depart- consideration. Applicants should send a curriculum vitae, publication list, ment of Geological and Environmental Sciences, Stanford University, synopsis of research interests and professional goals, and three letters of Stanford, CA 94305-2210. Questions can be directed to Prof. Donald reference (forwarded separately) to: Marine/Environmental Microbi- R. Lowe ([email protected]) or Prof. Stephan A. Graham ology Search Committee, c/o Ria del Rosario, Division of Biological ([email protected]). Sciences, 9500 Gilman Dr. 0366-A, La Jolla, CA 92093-0366. Stanford University has a strong institutional commitment to the principle of diversity. In that spirit, we particularly encourage UCSD is an Equal Opportunity-Affi rmative Action Employer applications from women, members of ethnic minorities, and with a strong institutional commitment to the achievement of individuals with disabilities. http://pangea.stanford.edu/. diversity among its faculty and staff. COLLEGE OF MEDICINE FACULTY POSITIONS

CENTER FOR IMMUNOLOGY

The Center For Immunology at the University of Illinois College of Medicine is inviting applications from individuals with a research focus in areas revolving around the central theme of immune tolerance such as Cancer Immunology, Persistent Infections, Cell/Organ Transplantation, Autoim- munity and Basic Immunology. The center will be housed in a new College of Medicine Research Building.

We are recruiting PhDs, MDs or individuals with an equivalent doctoral degree to fi ll a number of tenured / tenure track faculty positions at all academic ranks. Successful faculty candidates will be expected to have and maintain a vigorous independent research program as evidenced by extramural funding and consistent scholarly publications. Individuals will have primary appointments in an appropriate Basic Science or Clinical department. Generous laboratory space and start up funds are available.

For fullest consideration, please send applications, including CV, a brief statement of current research activities and future research plans and a list of 3 references to: Search Committee, Center for Immunology, University of Illinois College of Medicine, 835 S. Wolcott (MC 790), Chicago, IL 60612-7344.

For more information about the UIC Immunology Center, please visit our Web Site: http://www.uicimmunology.org.

VIROLOGY

The Department of Microbiology and Immunology in the College of Medicine at the University of Illinois at Chicago is inviting applications from individuals with a research focus in Virology revolving around viral replication, virus-host interaction, oncogenesis, pathogenesis and immunity. We are recruiting individuals with a PhD, MD or equivalent doctoral degree to tenured/tenure track faculty positions at all academic ranks. Successful faculty candidates are expected to have a vigorous research program as evidenced by extramural funding and consistent scholarly publications. The faculty of the Department of Microbiology and Immunology has active and interdisciplinary research programs spanning several major disciplines, including immunology, virology, microbial pathogenesis, host-pathogen interactions, and structural biology.

Please send applications with CV, a statement of proposed research program and a list of three references to: Virology Search Committee, Department of Microbiology and Immunology, University of Illinois at Chicago, College of Medicine, 835 S. Wolcott (M/C 790), Chicago, IL 60612-7344.

For more information about the Department of Microbiology and Immunology, please visit our Web Site: http://www.uic.edu/depts/mcmi/.

MICROBIAL PATHOGENESIS PROGRAM

The College of Medicine at the University of Illinois at Chicago is expanding its faculty in the area of Microbial Pathogenesis and is initiating an extramural search for full-time, tenured/tenure-track, faculty. Successful candidates will be appointed either in the Department of Microbiology & Immunology or the Divisions of Infectious Diseases or Gastroenterology in the Department of Medicine and most will have joint appointments. Investigators in these units along with faculty in the Department of Biological Sciences, the College of Dentistry, the College of Pharmacy and the School of Public Health provide a highly interactive research community.

We are seeking applicants at the ASSISTANT, ASSOCIATE or FULL PROFESSOR levels. Applicants must hold a PhD and/or MD degree with a proven track record in research as evidenced by consistent scholarly publications and extramural funding.

Please send applications with CV, a statement of proposed research program and a list of three references to: Dr. Linda J. Kenney, Chair, Search Committee, Department of Microbiology and Immunology, University of Illinois at Chicago, 835 S. Wolcott, (M/C 790), Chicago, IL 60612- 7344.

INFECTIOUS DISEASES AND IMMUNOLOGY

The Infectious Diseases Section in the Department of Medicine at the University of Illinois in Chicago is recruiting research faculty to fi ll tenure track positions. This recruitment is being done jointly with the Department of Microbiology-Immunology and Center of Immunology at UIC to expand multidisciplinary research between these programs. Faculty candidates will have MD, MD-PhD or PhD degrees and suffi cient experience to maintain independent, extramurally funded research programs. PhD candidates will have established extramural funding. ID Section faculty have existing programs that include studies of anthrax pathogenesis, viral oncogenesis, cryptococcal virulence and HIV pathogenesis and clinical trials. Joint initiatives with the Immunology Center include studies of viral oncogenesis and host response to infections.

For fullest consideration, please send an application, including CV, statement of research plans and 3 references to: Faculty Search Committee, ID Section (MC735), University of Illinois at Chicago, 808 S. Wood Street, Chicago, IL 60612.

For more information about the Section of Infectious Diseases, please visit our Web Site: http://www.uic.edu/com/dom/id/.

Review of applications for the above positions will begin April 1, 2005.

The University of Illinois at Chicago is an Affi rmative Action/Equal Opportunity Employer. Women and Minorities are strongly encouraged to apply. Science 2005. Assistant/Associate Professor Exhibit at February The Department of Materials Science and Engineering at MIT invites applications 21 for a tenure-track faculty position at the assistant/associate professor level, to begin the AAAS/ September 2005. Career Fair, Applicants should hold a Ph.D. in Materials Science and Engineering or a related science or engineering discipline. The successful candidate will be expected to develop a vibrant research program at the forefront of the field, and to harness their expertise in curriculum development and teaching at the undergraduate and graduate levels. Research areas of interest include, but are not limited to: materials chemistry, green Chemistry processes, sustainable growth technologies (such as energy production), combinatorial materials characterization, material interfaces, quantum processes in materials, soft materials modeling, clinical biomaterials, synthetic biology and nanostructured materials. Careers Applications should be submitted with two copies of the following: a complete A Science Advertising cv, statement of research and teaching interests, no more than three publications, Supplement and complete contact information for three references. Applications should be addressed to: Department of Materials Science and Engineering Att: Esther Greaves Estwick, Rm 8-328 Massachusetts Institute of Technology 77 Massachusetts Ave. Thomas Edison Cambridge, MA 02139-4307 1847–1931 Founder of Applications received by March 1, 2005 will receive full consideration. M.I.T. has a strong and Science continued commitment to diversity in engineering education, research and practice, and especially encourages applications from women and minorities.

Issue date web.mit.edu/hr 4 March 2005 Reserve ad space by 15 February 2005

Need to attract great scientists? The SystemsX initiative aims to place Switzerland among the leaders in systems Then talk to someone biology and functional genomics. As part of this initiative, the Faculty of Science who knows science. of the University of Zurich is seeking to fill the position of a non-tenured Bonus distributions: Assistant Professor in Systems Biology • American Chemical Society 13–17 March, San Diego, CA We are searching for outstanding individuals with a background in life sciences • / and a track record in the development of new approaches to systems biology and Northwestern Career Fair their application in the study of signal transduction, cell biology, or developmental 16 March, Chicago, IL biology. The successful candidate will be expected to establish an independent • Drug Discovery Europe research group within the Institute of Molecular Biology. The University of Zurich 14–16 March, London, UK will provide a competitive start-up package, state-of-the-art research facilities, and access to the Functional Genomics Center Zurich (www.fgcz.ch). There are excellent opportunities for interactions with other groups of the University of Zurich and the Swiss Federal Institute of Technology (ETH), as well as the National For more information, Centers for Competence in Research (www.snf.ch/en/rep/nat/nat_ccr_pro.asp). contact Daryl Anderson 202-326-6543 The position is initially limited to three years with the possibility of renewal for [email protected] another three years. Applications including a detailed curriculum vitae, publications list, short statement of research interests and the names and addresses of three referees should be submitted before March 31, 2005 to Prof. P. Truöl, Dean, Faculty of Science (MNF), University of Zurich, Winterthurerstrasse 190, CH- 8057 Zurich, Switzerland. The application material should also be submitted as a single Word- or PDF-file to [email protected].

The University of Zurich is an equal opportunity employer. Applications from women candidates are particularly encouraged. Department of Health        and Human Services National Institutes of Health      National Institute of Nursing Research            

                                     Scientifi c Director            Division of Intramural Research    With nation-wide responsibility for improving the              health and well being of all , the Department                 of Health and Human Services oversees the biomedical            research programs of the National Institutes of Health            and those of NIH’s research Institutes.            The National Institute of Nursing Research, a major             research component of the National Institutes of Health  !            (NIH) and the Department of Health and Human Ser-        " #      vices, is recruiting for a Scientifi c Director, a senior             scientifi c position to lead its intramural bio-behavioral     $ !       research program. The successful applicant will lead a developing group of independent principal investiga-             tors conducting research in the basic, behavioral and             clinical sciences relevant to the Institute’s mission (see               http://ninr.nih.gov/ninr/) and help chart the future of             ! a vibrant, growing nursing and bio-behavioral research              community across the NIH. The Scientifi c Director is expected to establish a cutting edge intramural program            of investigation into applying fundamental fi ndings to          %  bio-behavioral mechanisms. The successful candidate              will be responsible for allocation of the research budget              to intramural laboratory and clinical programs in coordi-       nation with rigorous, quadrennial reviews by the Board of Scientifi c Counselors. The Scientifi c Director will &      !     '  ( also be provided with a fully equipped new laboratory,              ! supported by resources that commensurate with the size    )*+         &  and scope of the program and will have the potential for       !,   ' '  ( recruitment of additional tenure-track investigators.         !  The successful candidate will have an earned doctor- -              ate in nursing or other health-related discipline, and an             established record of outstanding research accomplish- ments, management of research enterprises/programs,           interdisciplinary collaboration, and scientifi c leadership      .     / 0    and service within the nursing and/or bio-behavioral             1 community. Applicants should send a cover letter, cur-    2 riculum vitae, detailed statement of research interests and selected publications to: Dan Longo, MD, Co-Chair,                  NINR Search, c/o Dr. Melinda Tinkle, 31 Center                 Drive, MSC2178, Building 31/Room 5B-25, Bethesda,                  MD 20892-2178; [email protected]. Applications                  should be received by March 31, 2005.       DHHS and NIH are Equal Opportunity Employers.  Faculty Positions Stem Cell Biology University of California San Diego Division of Biological Sciences http://biology.ucsd.edu/ The Division of Biological Sciences at UCSD invites applications for Assistant, Associate, or Full Professor (tenure track or tenured) in stem cell biology, particularly from those working with human cells. All qualifi ed applicants are encouraged to apply, including underrepresented CLARK SCIENCE CENTER minorities and women. Instrumentation and Techniques Instructor Area of scholarship is open. We seek candidates to (Half-time, 3 year term, renewable) Center for Biochemistry complement our existing strengths in molecular, cell/developmental biology, and neuroscience, Qualifications: Ph.D. in Biochemistry or related discipline. In addition, at least one year of work experience and who will foster research involving stem cells. in biochemistry or related fields including proteomics is desirable. Broad expertise in biochemistry and in 2-D gel Applicants must hold a Ph.D. degree or equivalent electrophoresis and LC/MS is required; expertise in spectrophotometry, spectrofluorimetry, high pressure liquid and should have outstanding records of research chromatography, and/or capillary electrophoresis is desirable. achievement; above the rank of Assistant Profes- sor should have strong, extramurally supported Instrumentation and Techniques Instructor research programs. Appointees are expected to (3 year term, renewable) Center for Molecular Biosciences participate fully in departmental affairs/teaching. Appointment level will be commensurate with Qualifications: Ph.D. in Molecular Biology. In addition, at least one year of post-doctoral experience in molecular biology and bioinformatics including advanced DNA sequence and quantitative PCR analysis. Broad qualifi cations and experience with salary based expertise in most areas of molecular biology including genomic and cDNA library construction, PCR, Southern and on UC pay scales. Applications will be reviewed Northern blot analysis, DNA and RNA isolation, DNA fingerprinting, DNA sequence analysis, and quantitative PCR beginning March 15, 2005 and accepted until the and RT-PCR, and bioinformatics. Specific experience with Applied Biosystems instrumentation preferred. positions are fi lled. Review of resumes will begin immediately. Please forward resume, cover letter and three letters of Mail hard copies of curriculum vitae, publication recommendation, indicating position of interest to: I & T Instructors Search, Smith College, Clark Science list, synopsis of professional goals, research/teach- Center, Box 685, 117 Burton Hall, Northampton, MA 01063. ing interests, and three letters of reference (mailed directly from referees) to: Stem Cell Biology To view full job descriptions, please visit our website at: Search Committee, c/o Heather Pratt - 0376-B, www.smith.edu/hr/career_external.php University of California, San Diego, 9500 Gilman Drive, La Jolla, CA 92093-0376. UCSD is an Equal Opportunity-Affi rmative Action Employer with a strong institutional commitment to the achievement of diversity Smith College is an equal opportunity employer encouraging excellence through diversity. among its faculty and staff.

Medical University of South Carolina and the Hollings Cancer Center Director Institute of Cancer Drug Development This position is an exciting opportunity to develop a team of investigators who will explore new anticancer agents in the clinic, translating laboratory discoveries to the patient. The Institute will be housed in the new Hollings Cancer Center (HCC) laboratory space, the clinicians will be based in the Department of Medicine, and the basic researchers will have tenure track appointments in the basic science Departments in the School of Medicine. The successful applicant should have a signifi cant track record in develop- ing Phase I drug studies in the clinic, an independent clinical/basic research career with history of peer reviewed funding, and be a dynamic leader able to work within the Cancer Center matrix. The start-up package will include additional clinical recruitments and new laboratory space. The HCC will occupy a new building housing radiation therapy, outpatient clinics, and laboratory investigators. The Center has programs in Experimental Therapeutics, Hormone-Dependent Malignancies, Tobacco-related Malignan- cies, Functional Genetics, Cancer Biology and Prevention and Control. The Hematology/Oncology Division occupies an inpatient fl oor, sees patients in multidisciplinary clinics, and has an active bone marrow transplant program. The Medical University of South Carolina is located within minutes of the beaches, outstanding fi shing, the historic downtown and cultural venues, including the symphony, art, and history museums. Interested candidates should send a copy of their curriculum vitae and the names of three references to: Andrew S. Kraft, MD, Director Hollings Cancer Center, William H. Folk Chair in Experimental Oncology, 86 Jonathan Lucas Street, P.O. Box 250955, Charleston, S.C. 29425. Or Fax to 843-792-9456 or e-mail to Elisa Mundis at [email protected]. MUSC is an Equal Opportunity Employer, promoting workplace diversity. Tenure-Track Faculty Positions University of Texas, Brownsville

The Department of Biological Sciences announces four tenure-track faculty positions at the Assistant/Associate Professor level. The positions require expertise in: 1. Plant physiology, , crop biotechnology, or related areas. 2. Tropical ecology, coastal ecol- ogy, or marine biology. 3. Neurophysiology, neuropharmacology, or other neuroscience areas. 4. Infectious diseases, microbiology, or prokaryotic molecular biology. Qualifi cations include a Doctorate in a biological discipline and postdoctoral experience. Candidates will be expected to teach content-related courses. Applicants will be evaluated for their potential to develop an independent research program in their fi eld of expertise. Successful candidates are expected to begin their appointments in the fall 2005. The University of Texas at Brownsville is situated on the Southern tip of the continental United States, a distinctive and desirable sub- tropical location by the Gulf of Mexico and the US-Mexico border. Applications require a cover letter indicat- ing the area of expertise and qualifi cations, curriculum vitae, statements of teaching philosophy, research interests, transcripts and three references with contact information. The search will continue until the positions are fi lled. Submit your application to: Dr. Luis V. Colom ([email protected]), Biological Sciences, The University of Texas at Brownsville /Texas Southmost College (UTB/TSC), 80 Fort Brown, Brownsville, TX 78520. UTB/TSC is an Equal Opportunity Employer. Looking for a career that radiates success?

Then talk to someone who knows science.

If you want to shine in the world of science, it’s firmly founded on the expertise of Science,the essential you don’t leave your career to chance. At premier scientific journal, and the long experience ScienceCareers.org we know science. We are of AAAS in advancing science around the world. So committed to helping you find the right job, and to if you want a glowing career, trust the specialist in delivering the advice you need. Our knowledge is science. Visit ScienceCareers.org. Marie Curie 1867–1934 EPA’s Offi ce of Research and Development (ORD) is seek- The Minestry of Food, Agriculture ing candidates to fi ll approximately nine federal, four-year post- and Fisheries doctoral research positions. Recent initiatives at ORD facilities have promoted the conduct of cross-cutting research across the different The Danish Institute for ORD Labs and Centers in the areas of human environmental expo- Fisheries Research sure-effects and ecosystems. In the human health area, the overall mission for the cross-ORD post-docs will be to move forward more quickly the development and application of exposure, dose and Senior scientist health effects assessment methods or models. In the ecosystems research area, the cross-ORD post-docs will focus on advancing the in fisheries management modelling spatial analyses methods and on their application to water quality, The Danish Institute for Fisheries Research (DIFRES), ecological forecasting problems, and linkages between economic Department of Marine Fisheries, Copenhagen, Denmark, drivers and landscape conditions. Applications will be accepted invites applications for a permanent position as a senior from December 13, 2004 through March 31, 2005. scientist in modelling fisheries management systems. We are looking for a key research profile to participate in The duty station and organization will be based on the particular developing methodologies for and practical develop- disciplinary focus of the candidate and the overall requirements of ment of models/software for fisheries management the program. The positions will be in one of the following organiza- evaluations. The successful applicant is expected to tions: National Health & Environmental Effects Research Labora- strengthen the research profile of the fisheries section tory (NHEERL), National Center for Environmental Assessment ant to significantly contribute to the development of (NCEA), National Exposure Research Laboratory (NERL), and the the research area within the department. Place of employment: Copenhagen, Denmark. National Risk Management Research Laboratory (NRMRL). Duty stations associated with these organizations are: Research Triangle The full text of the announcement can be seen on Park, NC, Cincinnati, OH, Las Vegas, NV, Washington, DC, Athens, www.dfu.min.dk. Application deadline: 1st April 2005, GA, and Corvallis, OR. at 12.00 o’clock. Danish Institute for Fisheries Research (DIFRES) is a governmental These excepted service appointments offer a salary range of $48,947 research institution affiliated with the Ministry of Food, Agricultu- - $76,261 and include a full benefi ts package. Applicants must be re and Fisheries. The institution carries out research, investigati- US citizens or permanent residents. For specifi c job information and ons and provides advice concerning sustainable and quality- application instructions, you may access the ORD Internet site at: focused exploitation of live marine and fresh water resources. Thus DIFRES deals with chain considerations from water to table. http://www.epa.gov/ord/htm/jobs_ord.htm Danish Institute for Fisheries Research has approximately 280 employees located in Charlottenlund, Lyngby, Silkeborg and U.S. EPA is an Equal Opportunity Employer. Hirtshals.

New York University School of Medicine Faculty Positions in Medical and Molecular Parasitology

NYU School of Medicine is initiating a major expansion of the Depart-   ment of Medical and Molecular Parasitology under the leadership of our new Chair, Dr. Karen Day. We are seeking outstanding candidates to fi ll             four junior and senior level faculty positions.                              The Department of Medical and Molecular Parasitology aims to achieve               excellence in both research and teaching in the fi eld of parasitology.               We aim to discover and implement novel strategies to control parasitic             diseases, such as malaria, that affect the health of millions worldwide. The Department is playing a key role in new global health initiatives, including a Masters in Global Public Health, at NYU.              !                  Candidates working on host/parasite systems in any of the following   "       #$  $    research areas are encouraged to apply: Genomics, Epidemiology, !               % Genetics, Immunology, Biochemistry, Cell and Molecular Biology,              !      Anopheline Biology.                 Ideal candidates will complement and expand the existing strengths           in the Department. Candidates must have completed at least 5 years of postdoctoral training and developed an innovative research pro-        &    '  gram in the above mentioned research areas. Competitive startup &   (  )**+***  ,      ! packages and newly renovated laboratory space are augmented by          - .    $  core facilities including insectaries, bioinformatics, confocal micros- '  # /  01) #   %  ' 2  ' copy, animal facilities, flow cytometry. The Department is home 2  +**+* 3% (!- 4+45656)617 -  8   to 13 faculty with active parasitology research programs and an                   NIH-funded graduate training program (www.med.nyu.edu/ 9  +: ;  parasitology). See specific application instructions at http:// www.med.nyu.edu/parasitology/news/jobs.html. The School of Medicine was founded in 1841 and      is an Equal Opportunity, Affi rmative Action Employer. Women and minority candidates are especially encouraged to apply. Why not Change the World? BIOLOGY FACULTY SEARCH DEAN DEPARTMENT OF BIOLOGY School of Engineering and The Department of Biology at Rensselaer Polytechnic Institute seeks Applied Science candidates in any area of basic biomedical research for tenure track faculty positions at all academic levels. We are particularly interested in candidates in the areas of genomics, proteomics, and bioinformatics as part of a campus- The invites expressions of interest in, and wide initiative in computational biology and bioinformatics. nominations for, the position of Dean, School of Engineering and Applied Science. Rensselaer has recently opened a 218,000 sq. ft. Center for Biotechnology and Interdisciplinary Studies with approximately 60 faculty laboratories The University of Virginia is a vigorous, modern institution, and state of the art core facilities. Significant funding is available for animated by the forward-looking spirit of its founder, Thomas startup packages. Jefferson. In August 2004, U.S. News & World Report ranked the University of Virginia as the nation’s #2 public university (tied Review of applications will begin now, but the search will continue until with the University of Michigan), placing it 22nd overall among positions are filled. Please send a curriculum vitae, a statement of research 248 public and private colleges and universities. interests up to three pages, and have a minimum of three letters of reference sent to: The University's 2020 Commission on Science and Technology recommended three focused, multidisciplinary initiatives, which Robert E. Palazzo, Director, were launched in 2002: a University-wide information initiative to Center for Biotechnology and Interdisciplinary Studies, build excellence in computer and information science and in Biology 1W14 SC, Rensselaer Polytechnic Institute, engineering, an initiative in Quantum and Nanoscale Science and 110 8th Street, Troy, New York, 12180-3590 Engineering, and Morphogenesis and Regenerative Medicine. Contact Us at: Department of Biology, 1W14 Jonsson-Rowland Science The School of Engineering and Applied Science enrolls 2,645 Center, Rensselaer Polytechnic Institute, 110 Eighth Street, Troy, NY 12180- students, has an annual budget of $70 million, including $44 million 3590; Phone: (518) 276-6446. in externally funded research, a total full-time instructional/ research faculty of 167, and offers BS, ME, MS, and Ph.D. degrees. Rensselaer Polytechnic Institute is an Equal Opportunity, Affirmative Action Employer. As SEAS looks to its own future, four highly productive cluster Members of underrepresented groups (including people of color, or with disabilities, and women) are strongly encouraged to apply. areas have been identified for priority: bioengineering, computer and information science and engineering, nanotechnology, and societal and environmental systems. The individual selected for this prominent appointment will be a distinguished scholar with impeccable academic credentials, will have a deep appreciation for the research enterprise of a great university, and will be recognized as an accomplished academic leader. The Dean will have the breadth of view and experience needed to Case Western Reserve University School of Medicine appreciate and celebrate the multitude of research-intensive Endowed Chair in Infl ammatory Bowel Disease Research disciplines and educational programs both within SEAS and the broader University. The Dean should have a vision for opportunities Applications are invited from qualifi ed M.D., M.D.-Ph.D. or Ph.D. fac- in engineering that will inspire faculty, students, and administrators. A ulty to be appointed to the Victor and Ellen Cohn Chair for Infl ammatory high priority of this Deanship will be the ability to build new Bowel Disease at Case Western Reserve University/University Hospitals of relationships with external stakeholders and friends of SEAS and to Cleveland. The successful candidate will establish a cutting-edge research develop substantial new resources via capital fundraising initiatives. program to investigate the mechanisms and therapies of Crohn’s disease and ulcerative colitis. The Dean of SEAS will be an effective leader who will understand Applicants should have a strong academic track record commensurate and build upon U.Va.'s rich traditions and special resources as it with appointment as Professor or Associate Professor in the Department of continues as a premier institution. This person will take bold action Medicine (with tenure or tenure track) and an established research program and make decisions that reflect both intellect and foresight. This with sustained extramural funding in areas relevant to infl ammatory bowel leader will have a clear sense of vision based on strongly held disease, including infl ammation, gastrointestinal pathophysiology, genetics principles. The Dean will inspire others and instill pride in the and microbial pathogenesis, particularly if related to the enteric fl ora. School. The successful candidate will join and expand an internationally renowned This position represents an extraordinary opportunity for a group of investigators with an established track record of basic science renowned scholar and academic leader to be the Dean of investigation in infl ammatory bowel disease and mucosal immunity. In addi- Engineering of one of our country's premier universities. All tion to the salary provided by the Chair’s endowment, a generous package correspondence relating to the position of Dean of the School of commensurate with the applicant’s qualifi cations will be provided. Engineering and Applied Science should be directed in confidence Interested applicants should forward a letter of interest, a curriculum vitae, to the University’s executive recruitment consultant: the names and email addresses of three references, and a short summary of Dr. Randy Jayne and Karen Barrie their research plans to the Chairman of the Search Committee: Claudio Fiocchi, M.D, Division of Gastroenterology Heidrick & Struggles, Inc. Department of Medicine 1750 Tysons Blvd., Suite 300 University Hospitals of Cleveland/Case Western Reserve University McLean, VA 22102 (BRB 425), 10900 Euclid Avenue, Cleveland, Ohio 44106-4952 703-848-2500 Electronic format preferred to: [email protected] [email protected] Case Western Reserve University and University Hospitals of Cleveland are Affi rmative Action/Equal Opportunity Employers and applications The University of Virginia is an equal opportunity/affirmative from women and minority investigators are particularly encouraged. action employer committed to excellence through diversity. POSITIONS OPEN POSITIONS OPEN POSITIONS OPEN FACULTY POSITION EAST TENNESSEE STATE OPEN RANK UNIVERSITY Department of Cell Biology and Anatomy ASSISTANT or ASSOCIATE PROFESSOR of and the Stanley S. Scott Cancer Center Environmental Health. Two academic year tenure- Louisiana State University track positions beginning fall 2005. Minimum qual- Health Sciences Center SENIOR RESEARCH ASSOCIATE ifications include a graduate degree in environmental New Orleans policy, environmental management, industrial hy- A tenure-track faculty position at any rank is Pioneer Hi-Bred International, Inc is the world giene, occupational health, environmental health, or available in the Department of Cell Biology and leader in the discovery, development, and delivery of environmental engineering. Ph.D. and postdoctoral Anatomy and the Stanley S. Scott Cancer Center at elite crop genetics. We are looking for a Senior experience preferred. Candidates that are all but the Louisiana State University Health Sciences Research Associate for our Johnston, Iowa, location. dissertation will be considered. Must be able to teach Center (LSUHSC) in New Orleans. The successful This individual will provide antibody and enzyme- courses in one of the following areas: policy and law, applicant is expected to pursue a successful career that linked immunosorbent assay development support food sanitation and safety, solid and hazardous waste includes research and teaching excellence. This is a for crop genetics research and development. Respon- management, or occupational health and safety. joint recruitment involving the Department of Cell sibilities include monoclonal antibody screening, Ability to teach other courses in the Department is Biology and Anatomy and the Stanley S. Scott selection and evaluation, and developing and execut- desirable. The successful candidate(s) will demon- Cancer Center. The latter has partnered with the ing experimental strategies for assay development strate the ability to interface with a multidisciplinary Tulane University Cancer Center to form the and validation. Bachelor’s or Master’s Degree in faculty with a broad range of interests. Establishment Louisiana Cancer Research Consortium (website: biological sciences plus three years or more related of a funded research program in area of expertise that http://lacrc.net). Applicants must have a clear focus experience is required. Excellent communication involves graduate (Master_s) and undergraduate in cancer research, including active, or potential, skills are essential. students is expected. Competitive startup funds are funding by the National Cancer Institute, American The Req ID for this position is RP539. For a available. Applicants should send current curricu- Cancer Society, the Department of Defense, etc. complete job description and to apply, go to lum vitae and names, addresses, e-mail addresses, Areas of particular interest include cancer genetics and website: http://www.pioneer.com/employment. telephone numbers of at least three references to: DNA instability, molecular signaling (transcription Equal Opportunity Employer. Dr. Phillip Scheuerman, Chair, Department of factor, chromatin remodeling, cell-cell and cell matrix Environmental Health, College of Public and interaction, cell migration), and cell division, survival, BACTERIAL PATHOGENESIS Allied Health, East Tennessee State University, and apoptosis. Teaching responsibilities will be in the FACULTY POSITION Box 70682, Johnson City, TN 37614. Review of can- area of cell and molecular biology for health pro- Department of Medical Microbiology and didates will begin March 1, 2005, and continue until fessionals and graduate students. Immunology suitable candidates are found. An Equal Opportunity/ To apply, please send curriculum vitae, statement The Medical College of Ohio Affirmative Action Employer. of research interests, and the names of three The Department of Medical Microbiology and references to: Melissa Hebert, Business Manager, Immunology invites applications for an ASSIST- CENTER DIRECTOR Department of Cell Biology and Anatomy, Lou- ANT/ASSOCIATE PROFESSOR, tenure-track Richard B. Russell Research Center (RRC) isiana State University Health Sciences Center, faculty position in the area of bacterial pathogenesis. U.S. Department of Agriculture 1901 Perdido Street, Box P6-2, New Orleans, We are particularly interested in candidates focused Agricultural Research Service (ARS). The RRC is LA 70112-1393. on immune evasion and emerging/select bacterial located in Athens, Georgia, adjacent to the Univer- LSUHSC is an Equal Opportunity/Affirmative Action agents. Available facilities include a certified bio- sity of Georgia campus and includes offices, labo- Employer. safety level three laboratory and microarray/proteo- ratories, greenhouses, storage facilities, and 27 acres mics/tissue culture core facilities. We seek highly of land. The Center is responsible for the develop- ASSISTANT/ASSOCIATE motivated and collegial individuals with demonstrat- ment and dissemination of new technologies that PROFESSOR ed research productivity, a doctoral degree, and address a wide range of problems that impact the MICROBIOLOGY relevant postdoctoral training. The appointee will U.S. poultry industry and food safety issues impor- The Department of Biological Sciences at San Jose be expected to maintain a strong, extramurally tant to maintaining a safe food supply for the State University (SJSU) invites applications for a funded research program, participate in departmen- American consumer. The selectee will be responsible tenure-track position in microbiology at the Assist- tal teaching (particularly in the areas of bacterial for providing leadership, coordination, and vision ant/Associate Professor level to begin in fall 2005. genetics and pathogenesis), and interact productively for the Center, and administration of its resources. Applicants must possess a Ph.D. or equivalent with basic and clinical scientists focused on infec- Qualified candidates must have experience managing degree and be a broadly educated microbiologist tious diseases. Competitive salary, space, and startup a scientific/professional staff; skill in managing with a background in medical microbiology. Appli- funds will be provided. Applicants should send a brief multi-institutional programs; and knowledge of statement of research interests, representative pub- poultry and food safety research programs. U.S. cants should demonstrate a potential for excellence citizenship required. in teaching and an ability to establish a funded lications, curriculum vitae, and have three letters of Ph.D. preferred. ARS offers a research program appropriate for an undergraduate- recommendation forwarded to: Professor Garry T. comprehensive benefits package and negotiable re- oriented university. Such a research program is Cole, Medical College of Ohio, 3055 Arlington cruitment incentives, including relocation assistance. Avenue, Toledo, OH 43614-5806. Applicant Salary $82,438 to $126,064. For information con- expected to involve undergraduate and M.S. gradu- _ ate students. The candidate will coordinate an allied interviews will begin in March 2005 and will cerning the position contact: The Area Director s health microbiology course and teach in other continue until the position is filled. Visit our website: Office at telephone: 706-546-3311; for application upper-division microbiology courses and graduate http://www.mco.edu/depts/micro/search.html. information contact: Mrs. Genell Powers at tele- seminars. For consideration, send a letter of applica- Affirmative Action/Equal Opportunity Employer. phone: 706-546-3029. Applications must be post- tion, curriculum vitae, original university graduate marked by April 15, 2005. Visit our website: transcripts, separate statements of teaching and GEOBIOLOGY PROGRAM http://www.ars.usda.gov/careers/Careers.htm. research interests, and three letters of recommenda- University of Southern California USDA/ARS is an Equal Opportunity Provider and Employer. tion to: Chair, Microbiology Search Committee, The University of Southern California (USC) The Commodity Utilization Research Group, Department of Biological Sciences, San Jose State seeks applications for a tenure-track ASSISTANT/ Southern Regional Research Center, Agricultural University, One Washington Square, San Jose, ASSOCIATE PROFESSOR position in geobiol- Research Service, U.S. Department of Agriculture CA 95192-0100. Fax: 408-924-4840; telephone: ogy. We seek applicants with skills and interests in (USDA), New Orleans, Louisiana, is seeking a 408-924-4900. Review of applications will com- bridging the earth and biological sciences with novel permanent, RESEARCH , GS-1320- mence on February 18, 2005, and continue until the approaches and who will foster interdisciplinary 12/13. U.S. citizenship is required. Incumbent will position is filled. Starting date for the position can research and teaching. Review of applications will participate in a research project to utilize recombi- be August 2005 or January 2006. See website: begin in March 2005 and continue until the position nant DNA technology to engineer histidine acid http://www.sjsu.edu/depts/Biology for more is filled. Applications should include a complete phosphates for increased stability and specific activity information. SJSU is an Equal Opportunity/Affirmative resume, statement of research, and teaching experi- to hydrolyze phytic acid found in animal feed. Action Employer committed to the core values of inclusion, ence. At least three references should be sent directly Incumbent will work with a team of scientists to civility, and respect for each individual. to: Glen A. Smith (e-mail: [email protected]) and develop an enhanced phytase for animal feed be addressed to the committee chair. applications. A solid background in molecular ATMOSPHERIC SCIENTIST Dr. Kenneth H. Nealson modeling is required. Candidates must request a Responsible for weather risk management analyses c/o Glen A. Smith copy of vacancy announcement ARS-X5S-0083 by for insurance company. Ph.D. in physics, atmospheric Geobiology Search Committee Chair either calling telephone: 301-504-1482 or by sciences, or related field. Minimum one year experi- Allan Hancock Foundation copying the announcement from website: http:// ence including weather risk analysis, catastrophic risk AHF 107G, MC 0371 www.afm.ars.usda.gov/hrd/jobs/apply.htm in modeling, and pricing/loss assessment for insurance University of Southern California order to respond/submit specific information out- industry. Mail curriculum vitae to: P. Rademan, Los Angeles, CA 90089-0371 lined in the announcement. Applications must be ACE, 1601 Chestnut Street, TLP 33K, Philadel- USC is an Affirmative Action/Equal Opportunity postmarked by March 21, 2005. USDA is an Equal phia, PA 19103. Employer. Opportunity Provider and Employer. 784 4 FEBRUARY 2005 VOL 307 SCIENCE www.sciencecareers.org Department of Health and Human Services National Institutes of Health National Institute of Mental Health Molecular Imaging Branch Medical Staff Physician in Positron Emission Tomography The Faculty of Science (Mathematisch-naturwissenschaftliche Fakultät) of the The Molecular Imaging Branch at the National University of Zurich invites applications for a Institute of Mental Health, NIH has a medical staff physician position available on January 1, Professorship (Ordinarius, Extraordinarius, or Assistant 2005, with a flexible start date. This Branch has Professor with Tenure Track) in Systematic Botany embarked on major neuroimaging research to use state-of-the-art PET (positron emission at the Institute for Systematic Botany, to commence on October 1, 2007. The tomography) techniques and newly developed new professor is expected to establish and lead a successful research group, neuroreceptor radioligands to study neuropsy- contribute to the graduate teaching in Systematic Botany, and the undergraduate chiatric disorders. The NIH imaging facilities, teaching in Biology. She or he will find an excellent library, herbarium, laboratory radiochemistry laboratory, and multidisciplinary research team provide outstanding opportunities infrastructure, and botanical garden, as well as a stimulating environment. The for productivity. Imaging is performed in ani- institute is associated with the Zoological Museum, the Palaeontological Institute mals to evaluate new radioligand and in humans and other related research institutes in a School of Biology, and with other to study the pathophysiology of several neu- botanical institutes of the University, the ETH and the University of Basel in the ropsychiatric disorders. This senior position in Zurich-Basel Plant Science Center. Current research interests at the institute the laboratory will supervise trainee(s), fellow(s), include molecular and morphological phylogenetic systematics, comparative and support staff. The initial appointment is for evolutionary biology, and biogeography. 2 years, with option of unlimited renewals. Applicants may come from several medical We are looking for excellent researchers in systematic botany with an interest in specialties (e.g., Psychiatry, Neurology, or micro-evolution, palaeobotany, reproductive biology, morphology, or other fields Nuclear Medicine/Radiology) but must pos- that will complement existing research strengths in the institute. sess a valid US medical license and have several years of experience with radiotracer imaging. Applicants are invited to submit by April 30, 2005 a curriculum vitae, a publication Interested applicants should send CV and the list, and a summary of their research interests to Prof. Dr. P. Truöl, Dean of the names of two references to: Robert Innis, MD, Faculty of Science (MNF), University of Zurich, Winterthurerstrasse 190, PhD; Chief, Molecular Imaging Branch, NIMH, CH-8057 Zurich, Switzerland. The application material should also be submitted Building 1, Room B3-10, 1 Center Drive in a single Word or PDF file to [email protected]. MSC-0135, Bethesda, MD 20892-0135, Email: [email protected]. Women and minorities For further information, contact Prof. H.P. Linder, Institute for Systematic Botany, are encouraged to apply. phone +41 1 6358410, email: [email protected], or visit the institute DHHS and NIH are Equal homepage at www.systbot.unizh.ch. Opportunity Employers

The Mount Sinai School of Medicine seeks an outstanding professional who will be responsible Genomics Institute of the for leadership in research, teaching and the Novartis Research future growth of our Molecular Physiology and Department. Foundation

GNF focuses on paradigm-shifting technologies, making possible new Chair of Molecular approaches to complex biomedical problems. Pioneering technology platforms in genomics, proteomics, chemistry, structural biology, Physiology and Biophysics computation biology and engineering have been established as part of an integrated, multidisciplinary approach to biomedical research This position represents a unique opportunity to lead and expand a and drug discovery. Currently the institute consists of approximately department with exceptional research strengths in biophysics, 400 scientists dedicated to various areas of research including Neuro- biology, Cancer Biology, Immunology, Metabolic/Cardiovascular and structural biology, chemical biology and computational biology. We Infectious Disease. seek a candidate with outstanding scholarly achievements, a deep commitment to academic excellence, leadership skills, and a vision We are currently seeking the following positions: for basic science and translational research in an academic setting. Group Leaders, Embryonic Stem Cell Biology; Job Code MC4- 53 and Cancer Biology; Job Code GH3-89- We seek outstanding The school is particularly committed to expanding its research scientists to build research groups in stem cell and cancer biology. program in structural and chemical biology. The Chair needs to be We are especially interested in candidates who are able to utilize the committed to furthering interdisciplinary research, both within unique scientifi c environment of GNF to develop novel approaches to the basic biology of these systems. Candidates with 3-5 years of the department and in the larger institutional setting. He or she post-doctoral experience and an outstanding publication record are will also be expected to serve in a broader leadership role in the encouraged to apply. scientific enterprise within Mount Sinai School of Medicine. Visit our website: http://www.gnf.org for complete list of employment We offer competitive compensation and start-up package. Please opportunities. GNF offers excellent compensation and a great benefi ts forward your resume to: Dr. Dennis Charney, Dean of package. Mail to: 10675 John Jay Hopkins Dr., San Diego, CA 92121, Fax 858/812-1670, or submit online to [email protected] Resumes must Research, Box 1218, Mount Sinai School of Medicine, One include a Job Code for consideration. Gustave L. Levy Place, New York, NY 10029. Equal Equal Opportunity Employer. opportunity employer. POSITIONS OPEN POSITIONS OPEN POSITIONS OPEN DIRECTOR Program in Stem Cell Biology Yale University School of Medicine seeks an internationally The Donald Danforth Plant Science Center an- recognized academic leader to head a new Program TOXICOLOGIST nounces openings for PRINCIPAL INVESTIGA- in Stem Cell Biology. Qualified candidates must have The Science and Engineering Group of RTI TORS at advanced levels. We seek highly motivated strong research credentials and proven leadership International (Research Triangle Institute) is seeking scientists with established research programs that ability. This interdepartmental program will provide a candidate with a Ph.D. in toxicology to conduct complement those at the Danforth Center (website: a scientific focus for already strong efforts in research in proteomics as applied to toxicological http://www.danforthcenter.org). Of particular developmental biology, neurobiology, vascular biol- research problems. The individual will be responsible interest are candidates with interdisciplinary research ogy, hematology, immunology, transplantation, bio- for leading research projects in the Proteomics that targets plant biology as it impacts human and medical engineering, and translational research. We Research Group, working with existing and new animal nutrition/health, or production of bio-based encourage applications from individuals with in- clients, in both the commercial and government materials/products. The Danforth Center is set in terests in the fundamental aspects of mammalian sector, writing winning proposals, and collaborating suburban St. Louis, Missouri, and offers an stem cell biology, but candidates with other relevant with a highly interdisciplinary team of scientists in outstanding research environment with cutting-edge interests will also be considered. Qualifications biochemistry, analytical chemistry, proteomics, and core facilities in proteomics, microscopy, and cell include M.D., Ph.D., or M.D./Ph.D. degree. Can- bioinformatics. Strong communication and teaming imaging, bioinformatics, plant cell/tissue culture, didates are expected to meet the requirements for skills are necessary. and plant growth. Opportunity for adjunct academic appointment to the senior faculty ranks. Interested The applicant should have a Ph.D. degree in appointment at area universities. Salary will reflect candidates should electronically send curriculum toxicology and at least eight or more years of level of experience and prior record. Applicants vitae, brief statement of programmatic goals, and experience in the use of proteomic technologies as should submit curriculum vitae, selected reprints, names of references to e-mail: geraldine.emerling@ applied to toxicology directed studies. The candidate and a description of future research to: Ms. Billie yale.edu. should possess knowledge of routine and research Broeker, Human Resources Department, Donald Diane Krause, M.D., Ph.D. applications of modern proteomics technologies, Danforth Plant Science Center, 975 North Chair, Search Committee including two-dimensional gel electrophoresis and Warson Road, St. Louis, MO 63132. E-mail: c/o Gerri Emerling, Coordinator mass spectrometry. Experience shall include project [email protected]. Yale School of Medicine management, experimental design, data coordina- Equal Opportunity/Affirmative Action Employer/Minor- 333 Cedar Street, P.O. Box 208000 tion, and informatics in proteomic studies. The ities/Females/Persons with Disabilities/Veterans. New Haven, CT 06520-8000 candidate should also have experience in managing, mentoring, and directing the work of junior person- Yale University is an Equal Opportunity/Affirmative SCIENTIFIC DIRECTOR nel. A strong record in publishing articles in the Action Employer. Qualified women and minority group peer-reviewed literature and experience in preparing Center for Human Cell Therapy (CHCT) members are encouraged to apply. CBR Institute for Biomedical Research proposals to external funding agencies is mandatory. _ Please apply online at website: http://www. Children s Hospital, Boston CHIEF, PULMONARY UNIT. The Depart- Harvard Medical School rti.org/careers. Requisition No. CS11123. ment of Medicine of the We are proud to be an Equal Employment Opportunity/ The CBR Institute for Biomedical Research in- School of Medicine and Dentistry is recruiting for Affirmative Action Employer. Minorities/Females/Persons vites applications at the ASSISTANT or ASSOCI- the position of Chief, Pulmonary Unit. We are with Disabilities/Veterans. ATE PROFESSOR level for the position of looking for candidates with a strong record of Scientific Director of the Center for Human Cell independent research in pulmonary biology, admin- POPULATION BIOLOGIST. The Biology Therapy. The CHCT is a newly NIH-funded facility istrative leadership, and commitment to patient care Department of Franklin & Marshall College invites at the CBR Institute for Biomedical Research in the and education. Preference will be given to individuals applications for a one-year VISITING ASSIST- Longwood Medical Campus. The mandate of the with experience in the areas of inflammation and ANT PROFESSOR position in population biology CHCT is to provide resources to all Harvard immunology, pulmonary vascular disease, cancer, pending administrative approval, starting July 2005. Medical School faculty to facilitate bench-to-bedside and genetics. The successful candidate will be Candidates should have the Ph.D.; demonstrated development of cellular therapies–including thera- expected to foster the growth of innovative high strengths in teaching and research in both field and pies based on adult or embryonic stem cells–for the quality research and patient care programs in the laboratory settings; broad interests in population biol- treatment of damaged or diseased tissues. The Unit. Resources, including additional faculty po- ogy, evolution, and ecology; and strong quantitative Scientific Director of the CHCT will have close sitions, laboratory space in a newly constructed abilities. Teaching responsibilities will include lec- interaction with ongoing related research programs research building, office space, and startup funds tures and laboratories in an evolution-centered intro- at the Harvard affiliated hospitals and the Harvard (sufficient for four faculty recruits) are available to ductory course that includes Mendelian genetics and Stem Cell Institute. The incumbent of this position strengthen the highly successful Pulmonary Unit. ecology, and an upper-level elective in the candidate_s will be provided with startup funds and space to The successful candidate (M.D. or M.D., Ph.D.) area of expertise. Franklin & Marshall is a small direct his/her own extramurally funded research should have the academic qualifications for a tenure- (enrollment 1,800), highly selective coeducational program in areas related to stem cell or immuno- track appointment at the level of ASSOCIATE or liberal arts college with a tradition of excellence in therapy, such as stem/somatic cell biology, cell FULL PROFESSOR. The University of Rochester science and student research. Applicants should ar- trafficking, or transplantation biology. The incum- has undertaken a 10-year, $400 million strategic plan range to have letters sent from three referees, and bent will also provide oversight of the Translational to expand the Medical Center_s research programs. should submit curriculum vitae, plans for actively en- Cell Therapy Laboratory, which provides direction This recruitment is an integral component of the gaging undergraduates through teaching and re- to principal investigators engaged in translational strategic plan and a high priority. Interested appli- search, and undergraduate and graduate transcripts. research projects involving human cells. An M.D. cants should contact: Mark Taubman, M.D., Chief Priority will be given to completed applications re- and/or Ph.D. with evidence of outstanding investi- of Cardiology, 601 Elmwood Avenue, Box 679, ceived by March 18, 2004. Send applications to: gation and a record of collaborative interaction are Rochester, NY 14642. E-mail: mark_taubman@ Dr. Mark Olson, Department of Biology, Franklin essential. Prior experience overseeing a clinically urmc.rochester.edu. The University of Rochester is an & Marshall College, P.O. Box 3003, Lancaster, PA related resource, i.e., the development of human Equal Opportunity Employer and applications from women and 17604. Telephone: 717-291-4118; fax: 717-358- cells for therapy, is desirable but not required. To members of minority groups are encouraged. 4548; e-mail: [email protected]; website: apply, submit curriculum vitae, statement of research http://www.fandm.edu/biology.xml. The College is interests/plans, and names of three references via Bacterial Pathogenesis – Two POSTDOCTORAL committed to cultural pluralism through the hiring of minorities mail to: Leslie E. Silberstein, M.D., Director, POSITIONS are available: (1) to study the genetic and women. Equal Opportunity Employer/Affirmative Action. Joint Program in Transfusion Medicine, Chil- Francisella tularensis. The research will focus on dren_s Hospital Boston, Karp Family Research determining which genes from F. tularensis are One or two NIH-funded POSTDOCTORAL Building, 1 Blackfan Circle, 10th Floor, Room necessary for survival within macrophages and infec- POSITIONS available to study: (1) the role of 10217, Boston, MA 02115. tion of mice. This position requires select agent hepatocyte nuclear factor-4f C in redox regulation of Affirmative Action/Equal Opportunity Employer. clearance; (2) to study virulence regulation through hepatocyte inductible nitric oxide synthase gene quorum sensing in enterohemorrhagic E. coli. Can- transcription, (2) the role of nitric oxide in POSTDOCTORAL POSITION available in didates with a Ph.D. and a strong background in upregulation of osteopontin gene transcription, immunology, protein biochemistry, microbiology, molecular biology and prokaryotic gene regulation and/or (3) the effect of redox on chromatin re- or molecular biology to study novel innate immunity are highly desired. Please provide curriculum vitae modeling. M.D.-Ph.D. or Ph.D. candidates with a genes and their functions in transgenic mice, and the names and addresses of three references to: strong background in the molecular biology of tran- humans, and mammalian cells in vitro. Send curric- Dr. Vanessa Sperandio, Department of Microbiol- scription, protein-protein interactions, or protein- ulum vitae and names of references to: Dr. Roman ogy, U.T. Southwestern Medical Center, 6000 DNA interactions are encouraged to apply. Please Dziarski, Indiana University School of Medi- Harry Hines Boulevard, Dallas, TX 75390-9048. send curriculum vitae to: Paul C. Kuo, M.D., Duke cine,3400Broadway,Gary,IN46408,U.S.A. Fax: 214-648-5905; e-mail: vanessa.sperandio@ University Department of Surgery, Box 3522, Website: http://shaw.medlib.iupui.edu/nwcme/ utsouthwestern.edu. U.T. Southwestern Medical Center 110 Bell Building, Durham, NC 27710. E-mail: smpledsk.html. E-mail: [email protected]. is an Equal Opportunity/Affirmative Action Employer. [email protected]. 786 4 FEBRUARY 2005 VOL 307 SCIENCE www.sciencecareers.org GRANTS

CALL FOR GRANT APPLICATIONS Up to $500,000 per Award

Distinguished Scientist Award Three-year awards totaling up to $500,000

Through Distinguished Scientist Awards, The Sontag Foundation seeks to fund innovative brain cancer research while providing support and recognition to help advance the careers of outstanding brain tumor researchers who have recently attained an initial faculty appointment. These early career scientists must show exceptional promise of impacting brain cancer research through projects exhibiting the potential to lead to increased rates of survival and/or improved functional recovery for individuals with brain tumors.

Application and eligibility guidelines: www.sontagfoundation.com Application Deadline: March 22, 2005 Start of Award: On or about October 1, 2005

Eight outstanding researchers are currently being funded through The Sontag Foundation Distinguished Scientist Awards. Three additional awards are anticipated in 2005.

For further information please contact: The Sontag Foundation 822 A1A North, Suite 300 • Ponte Vedra Beach, FL 32082 904.273.8755 • email: [email protected]

The Science THE INTERNATIONALVACCINE INSTITUTE Meetings & POSITION ANNOUNCEMENT Announcements Institute Deputy Director for Laboratory Sciences The International Vaccine Institute (http://www.ivi.org) is seeking an Institute Deputy Director to Database oversee the operations and continue the development of the Institute’s Division of Laboratory Sciences, which will ultimately occupy approximately 90,000 sq feet of floor space in the Institute’s new headquarters facility. A comprehensive listing of The International Vaccine Institute is non-profit and focuses on research and development of new and events, grant announcements, improved vaccines for use in developing countries. The Institute’s headquarters are in Seoul, Korea. courses & training, and more … in print and online. The Laboratory Division currently has 7 principal investigator scientists working in laboratories devoted to mucosal immunology, cellular immunology, humoral immunology, bacterial genetics and bioinformatics, Your ad in Science is automa- and vaccine development and process research. The Division conducts both basic and applied research. tically posted in the Meetings Current laboratory work complements and takes advantage of the Institute’s ongoing epidemiologic and & Announcements database clinical research programs, which are being undertaken in 21 countries in Asia, Africa, and Latin America. at Sciencemeetings.org and receives a free hyperlink to The Division’s operating and research budget is supported by both grants and core funding. any e-mail or web address. While it is expected that the incumbent will possess a broad view of the entire field of vaccine research, he or Search this page by keyword, she should also be a recognized leader in a field relevant to the Institute’s laboratory research programs. It is discipline, geographic region, expected that the incumbent will conduct a program of independently funded research in his or her areas of or category/subject. It doesn’t interest, while maintaining overall responsibility for the Institute’s laboratory research programs. The incumbent get any easier. should also possess clear strengths in program-building, resource mobilization, and staff development. Minimum Is your event listed? qualifications include a doctorate degree in a relevant discipline, and significant experience in leading multi- person laboratory efforts in a research environment. U.S. – Daryl Anderson Salary will be internationally competitive. The Institute provides attractive fringe benefits including a 202-326-6543 housing allowance, home leave, and income tax reimbursement. Europe and International The International Vaccine Institute is an independent international organization established under the Tracy Holmes Vienna Convention of 1969. 35 countries and the World Health Organization are signatories to the +44 (0) 1223 326 500 Institute’s Charter. The Institute is governed by an international Board of Trustees, the majority of whom are elected based on their personal achievements and positions in the vaccine field. Letters of interest along with a current curriculum vitae should be addressed to: Mr. Hong-ki Jong, Senior Administrative Officer, International Vaccine Institute, Kwanak PO Box 14, Seoul, Korea. Tel: 82-2-872-2801 Fax: 82-2-872-2803 Email: [email protected] from whom further particulars can be obtained. Absolute confidentiality will be respected. POSITIONS OPEN POSITIONS OPEN POSITIONS OPEN TENURE-TRACK FACULTY The Albert Einstein College of Medicine seeks a POSITION tenure-track ASSISTANT/ASSOCIATE PRO- Immunoparasitology FESSOR in bioinformatics, who will serve as a The George Washington University Medical Cen- Director of the Bioinformatics Shared Resource ter is expanding its program in the Department of (BSR). The BSR is a data management/data mining Microbiology and Tropical Medicine (website: UNDERGRADUATE SUMMER facility, serving the research needs of the entire http://www.gwumc.edu/microbiology). We are RESEARCH OPPORTUNITIES medical school faculty. The successful applicant will now seeking applicants for a tenure-track position at During the 10-week (May 31–August 5, 2005), oversee a group of bioinformatics programmers, and the ASSISTANT PROFESSOR level whose re- NIH-supported program, students will conduct actively collaborate with basic and clinical science search focus is within the area of tropical and original research under the supervision of faculty researchers to provide data management and bio- infectious diseases, with special interest in immunol- members of the Department of Pharmacology. informatics analysis support. The position is highly ogy and vaccine development. The successful candi- Students will work on problems at the frontier of collaborative in nature and provides an opportunity date will be expected to establish an independent, biomedical receptor and signaling research using to establish an independent research program. The fully funded research program that will complement state-of-the-art techniques. In addition, students will BSR collaborates closely with statisticians, other the research activities of existing faculty in infectious participate in a foundations of signal transduction bioinformatics research faculty, and various AECOM diseases, vaccine biology, and immunology. We are course and workshops. At the end of the summer, shared resource facilities in proteomics, genetics, and particularly interested in individuals studying host- the students will present their work at a culminating clinical research. Screening of applications will be parasite interactions and their application to vaccine colloquium. continuous. Applications received by March 1, 2005, development and testing in animal models. Application deadline: March 1, 2005; salary: are assured full consideration. Required: A Ph.D. in microbiology, immunology, $5,000. Ref: Assistant/Associate Professor, Director of or infectious diseases with postdoctoral training is a Eligibility: chemistry, math, physics, computer Bioinformatics. prerequisite. Preferred qualifications: Applicants science, engineering majors who have completed Method of applying: Please send detailed curric- holding a D.V.M. or equivalent degree and previous their sophomore or junior years. U.S. citizen or ulum vitae with bibliography, and contact details experience in process development, vaccine formula- permanent resident status. (telephone number, e-mail address) for at least three tion, potency, and Quality Assurance/Quality Con- For application information: website: http:// references to e-mail: [email protected]. trol (QA/QC) are especially encouraged to apply. www.pharmacology.us/; Department of Pharma- Subject: Director of BSR; ATTN: Chair, Faculty The successful candidate will have an ongoing cology, School of Med- Search Committee for BSR sponsored research program, and some teaching icine, W1340 Biomedical Science Tower, Equal Opportunity Employer. experience. Pittsburgh, PA 15261. Telephone: 412-648- Review of applications will begin on March 20, 9321; fax: 412-648-1945; e-mail: summer@server. FACULTY POSITION 2005, and will continue until the position is filled. pharm.pitt.edu. IMMUNOLOGY/PATHOLOGY Submit curriculum vitae, a one- to two-page The University of Utah, Department of Patholo- statement outlining research and teaching interests, MICROSCOPY AND IMAGING gy, seeks qualified candidates in immunology for a and the names of three references to: CORE FACILITY MANAGER tenure-track position at the ASSISTANT/ASSO- Peter Hotez, M.D., Ph.D. The Neurobiology Program of Children_s Hospi- CIATE PROFESSOR level, with emphasis on host Department Microbiology and Tropical Medicine tal Boston, Harvard Medical School, seeks a System responses to bacterial, viral, or parasitic infections. The George Washington University Manager for the Imaging Core Facility of the NIH- Candidate should have a Ph.D. and/or M.D., and Ross Hall, Room 736 funded Mental Retardation Research Center. The a record of scholarly, laboratory-based achievements 2300 Eye Street, N.W. Core serves as a resource for single and multi- with strong potential for independent, extramurally Washington, DC 20037 photon laser scanning confocal microscopy, digital funded research. Telephone: 202-994-3532 microscopy, and advanced image analysis to inves- Curriculum vitae and brief statement of current Fax: 202-994-2913 tigators throughout Children_s Hospital and Har- and future research should be submitted by April 1, E-mail: [email protected] vard Medical School. Applicants should be 2005. Please include the names of three individuals The George Washington University is an Equal Opportu- knowledgeable in the use and maintenance of who can provide recommendation letters. nity/Affirmative Action Employer. confocal and standard microscopes, and image c/o Janis Weis, Ph.D. analysis software programs. Responsibilities will Department of Pathology POSTDOCTORAL POSITION. Kidney Biol- include basic Core administration, assisting inves- University of Utah ogy and Pathobiology, Case School of Medicine tigators with use of microscopy equipment, and the School of Medicine (website: http://www.metrohealthresearch.org/ design of new imaging techniques, and maintaining 30 N 1900 East Room 5C124 renal.html). Unexpected opening on NIH institu- the Core instruments. Preference will be given to Salt Lake City, UT 84132 tional training grant beginning July 2005. Training candidates with training or experience in multi-label Website: http://www.path.utah.edu/cbi/index. faculty use cell, molecular biology, and genetic intracellular localization, fluorescence recovery after htm. techniques to understand normal kidney function photobleaching, fluorescence resonance energy The University of Utah is an Equal Opportunity/ and mechanisms of kidney disease. Research in- transfer, and tissue slice, live cell, and whole mount Affirmative Action Employer. terests include identification of nephropathy suscep- imaging. Good communication skills are essential. tibility genes, expression profiling at an RNA and Minimum requirements include a Bachelor_sor protein levels, analysis of transcriptional regulation, Master_s degree in a related field with microscopy ex- ASSOCIATE RESEARCH regulation and function of novel kidney ion chan- perience. Send resume and two letters of reference to: SCIENTIST nels and transporters, control of protein trafficking, Scott L. Pomeroy, M.D., Ph.D. Institute for podocyte biology, extracellular matrix receptors and Imaging Core Director Cancer Genetics signaling, kidney development, effects of mechanical Mental Retardation Research Center Associate Research Scientist positions are availa- force on cells, and mucosal immunity. M.D. or Neurobiology Program ble to study the molecular pathogenesis of various Ph.D. candidates are welcome. Experience in mo- Enders 260 cancers, including breast, lymphoid, prostate, and lecular biology, cell biology, biochemistry, or elec- Children_s Hospital Boston brain. Ph.D. or M.D. and extensive related research trophysiology is desirable. Must be permanent 300 Longwood Avenue experience required. resident. Send letter of application, curriculum vitae, Boston, MA 02115 Please send statement of research interests, cur- and letters of recommendation, c/o e-mail: nkessler@ E-mail: [email protected] riculum vitae, availability, and names of references metrohealth.org. to: Dr. Linda Lowenstein, Institute for Cancer POSTDOCTORAL POSITIONS Genetics, Columbia University, 1150 St. Nicholas POSTDOCTORAL FELLOW Avenue, New York, NY 10032. Fax: 212-851-5256 Two NIH-funded positions are available in the or POSITION e-mail: [email protected]. area of flavoprotein enzymology in the laboratory of Columbia University is an Equal Opportunity/Affirmative A Postdoctoral position is available to investigate Dr. Marilyn Jorns at College of Action Employer. flagellin glycosylation and the role of flagella in Medicine, Department of Biochemistry and Molec- virulence of . The position is ular Biology, Philadelphia, Pennsylvania. These located in the laboratory of Dr. Patricia Guerry, positions involve mechanistic studies on bacterial POSTDOCTORAL POSITION Enteric Diseases Department of the Naval Medical sarcosine oxidases and nikD, an enzyme important in University of California, San Francisco Research Center in Silver Spring, Maryland, a the biosynthesis of nikkomycin antibiotics (website: Available to study neuronal circuitry underlying suburb of Washington, D.C. Candidates must have http://www.drexel.edu/med/biochemistry/faculty/ behavior in zebrafish. Candidates should have a a Ph.D. degree in microbiology, molecular genetics, resume_jorns.htm). A general background in strong background in molecular biology, micro- chemistry, or related discipline. Please send applica- biochemistry with experience in mutagenesis/ scopic imaging, and neurobiology. Please send curric- tions, including detailed curriculum vitae and con- kinetics and molecular biology is desirable. Please ulum vitae and names of three references to: Dr. Su tact information for three references to website: forward curriculum vitae and the names/contact Guo, Department of Biopharm. Sciences, Univer- http://www.hjf.org. Attn: Job Req. 200973. Affir- information of three references to e-mail: marilyn. sity of California, San Francisco, CA 94143-2911. mative Action/Equal Employment Opportunity. [email protected]. E-mail: [email protected]. 788 4 FEBRUARY 2005 VOL 307 SCIENCE www.sciencecareers.org 3.#46 21/+6,36$+,36+836 //8 217

638/ 2 %374%36+8 32 6QRKEU KPENWFG ; 2+7/ %374%+8+7/ +8%;+7 2/8+,3 7 /# 12+ 21 /-/ + 789  ; 2+7/ %374%+8+7/,236+ 8 /7 2/9+2 7/+7/ ,786+-879//,69+6 92//# 12+ 21 837-//832+2./.%/7 32# 12+ 21 1TICPK\GTU # 12+ 21,"/-/4836; 2+7/7 %374%+8+7/7 #+6+39682/ .1/,&+22./"/7/+6-9278898/ /9632+# 12+ 21 /2:+ 2//,/8976+//+-32/77/.-+/28/6  --$32-7,3.#4621-+6,36:+,36+836 5GUUKQP %JCKTU "3,/68,6+%+,962+ 9278898/ -G[PQVG #FFTGUUGU +02++68#+1 ,#832633>%2:/678 6+ 1$%34732,%2:/67830 /227:+2+ 36//2+286/,%2:/6783092.// +68 2/"3977/,#8 9./.6/2 7"/7/+6--3748+ <+-- "32,%#+2/13 ;+8%39.,--9 &+2./6,8%2:/678  9/6,-%2:/678

8%/6 //8 217 $%/ 33130/23/7  -63, + +8%31/2/7 7 $+61/8/0 2 8 32&/-836/7 12 +   /378"/74327/ 0363/-9+6/. - 2/ "/863: 697/7 #/48/ ,/6   +6- + 6316+/.//+8% +1 21/96327/96+-8 : 8  8%#437 93/-9+6 #/48/ ,/6  +6-   4463+-%/78332863 21+2-/6 " #78/7 3313,+"/19+8 32 92/  #/48/ ,/6 -83,/6 30/2/"46/77 32 ,/+78/ 331 /963, 33130&TQUQRJKNC +6-  91978  -83,/6 $/+62 21/36 9-+638 -" 63-/77 21 /23/2036+8 -7 46  91978 -83,/6 $%/%, 59 8 2+  /-%+2 77309-+638 - 3/-9+64463+-%/783 46+ $6+27-6 48 32 &+-- 2//7 12 $/3/6/7$/3/6+7/ 91978 #/48/ ,/6 /-/ ,/6  + 9-+638 -"/4 -+8 32 "+8/23 -73./7 #/48/ ,/6 /-/ ,/6 

3.#46 21/+6,36$+,36+836e//82173967/7 6316+ e 92183;2"3+.,3.#4621-+6,36,/  ;;; -7% /.9 //8 217 e432/01 2e/ +0 //8217-7 /.9e0+601 2 POSITIONS OPEN POSITIONS OPEN POSITIONS OPEN COMPUTATIONAL SCIENTISTS POSTDOCTORAL AND CLINICAL POSTDOCTORAL RESEARCH Gene Expression Biostatistics and POSTDOCTORALFELLOWSHIPS AND CLINICAL FELLOW Data Integration FELLOWSHIPSat the The Division of Rheumatology at The University Computational scientists will be responsible for National Institutesat the of Health of Texas Southwestern Medical Center at Dallas is analysis of microarray data, integration of the results NationalU.S. Department Institutes of of Health Health seeking a Postdoctoral Research Fellow with experi- with other proprietary and publicly available data, U.S.and Department Human Services of Health ence in the area of immunology and inflammation. design and implementation of high throughput sys- Website: http://www.training.nih.govand Human Services The successful candidate must have a Ph.D. or tems for genomic and metabolic pathway discov- Website:NIH ishttp://www.training.nih.gov dedicated to building a diverse equivalent in immunology, nephrology, or related ery, and development of data mining interfaces. communityNIH is dedicated in its training to building and employment a diverse disciplines and is expected to engage in externally The candidates should be highly motivated, have a community in itsprograms. training and employment sponsored research programs in lymphocyte and Ph.D. or M.Sc. in statistics, computer science, phys- programs. responses to oxidative stress and auto- ics, or biology, a solid background in statistics and immune inflammation, present data, and to publish algorithm development, extensive experience of ex- in peer-reviewed journals. Interested candidates perimental data analysis, and proficiency in pro- FACULTY POSITION should submit a letter of application designating gramming. Experience with commercial data analysis The University of Chicago current and long-term research interest, curriculum software Genespring in desirable. They also should Gwen Knapp Center for Lupus and vitae, and the names and addresses of three refer- have at least four years of software development ex- Immunology Research ences to: perience in UNIX or Microsoft environments, with The Gwen Knapp Center for Lupus and Im- David R. Karp, M.D., Ph.D. proven experience in Java, HTML, JSP, and web in- munology Research at the University of Chicago Associate Professor of Internal Medicine terface development. The individuals must be profi- is seeking a tenured position in immunology. This Chief, Rheumatic Diseases Division cient in SQL, PL/SQL, and UNIX shell scripting, be appointmentispartofanexcitingandrenewed UT Southwestern Medical Center familiar with database design, and possess excellent expansion of the Center and the Department of 5323 Harry Hines Boulevard communication skills. Experience in the following Pathology. Dallas, TX 75390-8884 areas is a plus: Visual Basic, XML, and PERL, J2EE Successful applicants should hold an M.D., Ph.D., E-mail: [email protected] and application frameworks (Spring, Webwork, etc.). or M.D./Ph.D. and have an outstanding record UT Southwestern Medical Center is an Equal Opportuni- Successful candidates will be able to work closely with of published research and sustained peer-reviewed ty/Affirmative Action Employer. staff scientists on different biological projects. Please funding. The academic rank will be commensurate refer to Job Code Bi-0105. with qualifications and will be in the Department Ceres, Inc. has moved to a new state-of-the-art of Pathology. Applicants must have a strong back- ASSOCIATE RESEARCH SCIENTIST and/ research facility in the hills of Thousand Oaks. We ground in lupus research. A commitment to educa- or POSTDOCTORAL RESEARCH SCIENTIST offer competitive salaries and excellent benefits in- tion (residency training and/or graduate or medical position available immediately in the Department of cluding equity participation and a 401K. Highly student training) is essential. The Knapp Center of- Pharmacology at Columbia University. This is an motivated individuals should send applications, in- fers excellent laboratory facilities and access to out- opportunity to join an interdisciplinary team inves- cluding current curriculum vitae, summary of re- standing core facilities at the University of Chicago tigating the molecular basis of inherited mutation- search experience, and names and addresses of three including flow cytometry, confocal/immunocelectron induced rhythm disturbances in the heart. Candidates references to: Human Resources Manager, Ceres, microscopy, DNA and peptide synthesis, microarray must have either Ph.D. or M.D. degrees and must Inc., 1535 Rancho Conejo Boulevard, Thousand facility, and immunohistochemistry. A highly com- have experience appropriate for the positions. Exper- Oaks, CA 91320; or e-mail to: ceres-hr@ceres-inc. petitive startup package is available. tise in protein chemistry and/or biochemistry of mem- com. The positions are available immediately. Applicants should submit by, March 1, 2005, their brane and signaling molecule proteins is required. Department of Health and Human Services curriculum vitae, a short research summary with plans Send curriculum vitae to: Dr.RobertS.Kass,Ph.D., National Institutes of Health for further research, and three letters of recommen- Department of Pharmacology, Columbia Univer- National Cancer Institute dation to: Martin Weigert, Ph.D., Director, Gwen sity College of Physicians and Surgeons, 630 West Knapp Center for Lupus and Immunology Two POSTDOCTORAL POSITIONS are 168th Street, New York, NY 10032. Research, The University of Chicago, 924 East We are an Equal Opportunity/Affirmative Action Institution. available with the Laboratory of Protein Dynamics 57th, R413A, Chicago, IL 60637. E-mail: and Signaling, Center for Cancer Research (CCR), [email protected]. National Cancer Institute (NCI), National Institutes The University of Chicago is an Affirmative Action/Equal of Health (NIH), Department of Health and POSTDOCTORAL FELLOWSHIP position is Opportunity Employer. Women and minorities are encouraged available at Northwestern University, to work in Human Services (DHHS) located in Frederick, to apply. Maryland, to: (1) study the role of the ubiquitin projects on the mechanisms of interferon and system in Parkinson_s Disease and (2) BOSTON HARBOR ISLANDS retinoic acid signaling in malignant cells. Previous study structure/function relations of ubiquitin POSTDOCTORAL FELLOWSHIP experience in molecular biology/biochemistry ligases and participate in the development of small The Department of Entomology in the Museum preferred. Applicants should submit their curriculum molecules that modulate the activity of ubiquitin of Comparative Zoology (MCZ) invites applications vitae to: Leonidas C. Platanias, M.D., Ph.D., ligases that are implicated in cancer. Background in for the Boston Harbor Islands Postdoctoral Fellow- Professor of Medicine, Deputy Director, Robert cancer cell biology, biochemistry, or neurobiology is ship, located in the existing biodiversity and island H. Lurie Comprehensive Cancer Center, North- highly desirable. Candidates should have a recent biogeography program in the MCZ. The incumbent western University, Feinberg School of Medicine, M.D. and/or Ph.D. and less than five years of will work with MCZ and National Park Service Olson 8250, 303 East Chicago Avenue, Chicago, postdoctoral experience. (NPS) personnel to develop an All Taxa Biotic IL 60611. E-mail: [email protected]. Salary is commensurate with research experience. Inventory (ATBI) with emphasis on insects and Please apply electronically by sending letter of other invertebrates located on the Boston Harbor interest, curriculum vitae, and names of three refer- Islands national park area. Additional expectations The Plant Science Research Center at the Uni- ences to: Allan M. Weissman at e-mail: amw@ include participation in educational outreach; for versity of Toledo has two openings for POST- nih.gov. example, assisting in the development of an online DOCTORAL RESEARCH ASSOCIATES, three DHHS and NIH are Equal Opportunity Employers. database of the ATBI ongoing results and linking openings for RESEARCH LABORATORY TECH- them to websites: http://insects.oeb.harvard.edu NICIANS, and one opening for a GREENHOUSE POSTDOCTORAL POSITIONS are available and http://www.bostonislands.org, and in the TECHNICIAN. Please visit website: http://psrc. to examine the structure and function of porins of development of a public exhibit for the Harvard utoledo.edu for position descriptions and further Mycobacterium tuberculosis. The unique outer Museum of Natural History using this data information. Review of applications will begin from membrane of Mtb is an efficient permeability obtained. The Fellowship is available for three years, February 14, 2005. The positions will remain open barrier that is the major determinant of the intrinsic with a desired start date of May 1, 2005, and until they are filled. The University of Toledo is an Equal resistance of mycobacteria to most antibiotics. The possibility of renewal for two additional years. Please Access/Equal Opportunity/Affirmative Action Employer and ultimate goal of this NIH-funded project is to send a cover letter with a statement of relevant Educator. exploit this knowledge for the design of more experience and intent, curriculum vitae, representa- effective TB drugs. For further information see: tive publications, and arrange to have three letters of ANNOUNCEMENTS Mol. Microbiol. 49:1167, 2003; Science 303:1189, reference sent to: 2004. UNIVERSITY OF CAMBRIDGE Supplementation to NIH salary levels is available Committee for Boston Harbor Science Summer School for qualified candidates. The new laboratory, the Postdoctoral Fellowship 17 July–6 August 2005 Department of Microbiology, and the University of Department of Entomology Study with some of Cambridge"s finest scientists Alabama at Birmingham provide an outstanding Museum of Comparative Zoology and other leading experts. Choose from a broad research environment. The establishment of inde- 26 Oxford Street range of science topics to suit your particular inter- pendent projects is encouraged. Please send curric- Cambridge, MA 02138 ests. Further information at website: http://www. ulum vitae and three letters of recommendation to: Review of applications will begin on March 15, cont-ed.cam.ac.uk/IntSummer/sschools/Science/. Michael Niederweis, Ph.D., e-mail: mnieder@ 2005. Harvard University is an Equal Opportunity E-mail: [email protected]. Telephone: uab.edu. UAB is an Equal Opportunity Institution. Employer. þ44 (0)1954 280398 (quote ref:SCI). 790 4 FEBRUARY 2005 VOL 307 SCIENCE www.sciencecareers.org NIH-RAID (Rapid Access to Intervention Development) AN NIH ROADMAP INITIATIVE

Consultants to the Roadmap planning process advised the NIH to facilitate the development and testing of novel therapeutic interventions, particularly ones for uncommon disorders which may not attract private sector investment. Based on this advice, the NIH is establishing a new program, NIH-RAID, modeled upon the NCI’s RAID program, to make available on a competitive basis, contractual resources for the preclinical 14-17 March 2005 development of small molecules. Novotel London West, UK NIH-RAID is not a grant program. Instead it will afford investigators access to critical resources needed for preclinical development. Services may include: creation of a product development plan, small molecule • Distinct, original scientific and synthesis according to GMP standards, formulation, pharmacokinetic business content focusing on studies and animal toxicology, and help with preparation of an IND. R&D productivity presented by Services actually provided will depend upon the stage of the project and the strength of the preliminary data. The fi nal product will be returned to an international panel of the applicant for further target validation and/or phase I human studies. industry experts The deadline for receipt of requests is June 1, 2005. • Drug discovery case histories • Further information about this program and the required format for dedicated to providing concrete applications can be found at: http://nihroadmap.nih.gov/raid/. examples of the latest product • Inquiries can be made to the NIH-RAID Program Coordinator by telephone at 301-496-6325 or by e-mail to [email protected]. development trends NIH-RAID PILOT Program Offi ce NIDDK For further details please visit: 6707 Democracy Blvd National Institute of 2 Democracy Plaza, Room 634 www.drugdisc.com/europe Diabetes & Digestive & Kidney Diseases Bethesda, MD 20892-5460

Telephone (301) 594-4660 New for 2005 Co-Located with: [email protected]

The leading provider of scientific, technological and business information

Science 2005. Exhibit at 21 February the AAAS/ Career Fair, Women

in Science meetings and announcements Advertising Supplement

Issue date 25 February 2005 Reserve ad space by 8 February 2005

Bonus distributions: • Association for Women in Science • American Men and Women in Science mailing list • University of Wisconsin Career Fair 2 March, Madison, WI For more information,contact: Daryl Anderson – 202-326-6543 [email protected] AAAS Annual Meeting and Science Career Fair

The 2005 AAAS Annual Meeting is the perfect place to explore your career options and attend the Science Career Fair. Both the career fair and the career-related workshops are FREE to attend.

AAAS Annual Meeting AAAS/Science Career Fair

DATES: 17–21 February 2005 DATE: 21 February 2005

PLACE: Marriott Wardman PLACE: Marriott Wardman Park Hotel Park Hotel Washington, DC Washington, DC

TIME: 11:00 am – 4:00 pm

CAREER WORKSHOPS (see website Science Careers offers you the for complete listing of workshops ): chance to meet employers. • Strategic Networking Exhibiting employers are typically from biotechnology, pharmaceutical, • Pathways to Multiple Career Opportunities government, and manufacturing organizations. • AAAS Fellowship Program in Public Policy and Mass Media For more information and updates to our exhibitor list, please visit • Research Training at the NIH www.sciencecareers.org and click meetings and announcements • How to Fire Up your Presentation on Career Fairs.

Registration is required to M attend the career workshops. Visit www.sciencecareers.org and click on Career Fairs for instructions on how to register for free. 2005 FASEB SUMMER RESEARCH CONFERENCES

Saxtons River, Vermont  Autoimmunity  Mechanisms & Regulation of Prokaryotic Transcription  Nuclear Structure&Cancer – Co-sponsored by The Endocrine Society  Proteases in Hemostasis & Vascular Biology  Transport ATPases: Genomics, Mechanisms,&Relevance to Diseases  Perspectives in Transport Biology  Hematological Malignancies  New Insights in Polycystic Kidney Diseases: Molecular Pathways, Pathogenic Mechanisms, & Translational Applications

TUCSON, ARIZONA  Mammalian Mobile Elements  Skeletal Muscle&Stem Cells  Biology & Chemistry of Vision  Intracellular RNA Sorting, Transport & Localization  Lymphocytes & the Immune System: Molecular, Cellular & Integrative Mechanisms  Calcium Oxalate in Biological Systems  Immunoreceptors  Nutrient Control of Gene Expression & Signaling – Co-sponsored by The Endocrine Society  Growth Factor Receptor Tyrosine Kinases in Mitogenesis, Morphogenesis & Tumorigenesis

SNOWMASS VILLAGE, COLORADO  Regulation of Ion Channel  Lysophospholipid Mediators in Health and Disease  Signal Transduction in the Immune System  TGF-ß Superfamily: Signaling & Development  Chromatin & Transcription  Protein Kinases & Protein Phosphorylation

 & Genome Rearrangements meetings and announcements  Receptor & Signal Transduction – Co-sponsored by The Endocrine Society  Glucose Transporter Biology – Co-sponsored by The Endocrine Society  Gastrointestinal Tract XI: Innovations in GIResearch & Therapy

TOSCANA, ITALY  Ciliate Molecular Biology

Mailing List To be added toour mailing list,contact: FASEB Summer Research Conferences FAX: 301-634-7007 [email protected]; [email protected];[email protected]

Information & Application Available in March on our web site –http://src.faseb.org FASEB Members will Automatically Receive the Meeting Notice MARKETPLACE MARKETPLACE

%WUVQO 2GRVKFGU #PVKDQFKGU $GUV 5GTXKEG2TKEG%QORCTG CPF 5CXG Who’s cultivating tomorrow’s (TGG 5GSWGPEG CPF #PVKIGPKEKV[ #PCN[UGU #NRJC &KCIPQUVKE   scientific geniuses? YYYCFKEQO service"CFKEQO

Ava att P1 Pst I Diverse Small Molecules Apa LI Apa LI Ava I Simulate Xma I i C x t r c loning e perimen s o Sma I Ready for Scree i c n ng d B Upwards of 200,000 ChemBr e pDonor 201 Draw idg d 0 b c Pub cat Qua ty a Compounds Corporat on 447 p c li ion li m ps i K m Bam HI r r m Pre-Plated in DMSO C S mVector Eco Very Competitively Pst attP2 Priced www.PremierBiosoft.com 650-856-2703 Website: www.chembridge.com Next Day Delivery* Email: [email protected] (800) 964-6143 or (858) 451-7400 Fax: (858) 451-7401 * Limited to 100,000 POLYCLONAL ANTIBODIES Lets Us Design Your …MADE EASY! Antigen for FREE! The World of Science Online FAST DELIVERY PEPTIDE TO ANTISERUM IN 70 DAYS SAGE KE 100% SATISFACTION GUARANTEED Tel: 888-343-5974 E-Marketplace Fax: 978-630-0021 www.newenglandpeptide.com ScienceCareers.org Questions and Answers. Science’s Next Wave www.scienceonline.org Some particularly gifted children might be able to Science NOW STKE make quantum leaps in their education and find science a relatively easy subject to comprehend. Others may need a little more help and encouragement at an early age. Helping develop that interest and provide the learning tools necessary is something we High throughput DNA sequencing at AAAS care passionately about. It’s a big part of the Gene synthesis $2/bp any size very reason we exist. Protein expression & purification Our educational programs provide after-school Yeast 2 hybrid/phage displaying activities such as the Kinetic City web-based science www.mclab.com, 888-625-2288 adventure game, based on the Peabody Award winning Kinetic City radio show; Science Netlinks, with over 400 science lessons available on the Internet; and Project 2061, which provides teaching benchmarks to foster an improved understanding of science and technology in K-12 classrooms. AAAS has been helping to answer the questions of science and scientists since 1848, and today is the world’s largest multidisciplinary, nonprofit membership association for science related professionals. We work Looking for a hard at advancing science and serving society – by supporting improved science education, sound science policy, and international cooperation. job? So, if your question is how do I become a member, here’s the answer. Simply go to our website at • Job Postings www.aaas.org/join, or in the U.S. call 202 326 6417, or internationally call +44 (0) 1223 326 515. • Job Alerts Join AAAS today and you’ll discover the answers • Resume/CV are all on the inside. Database • Career Advice • Career Forum

www.aaas.org/join

794 4 FEBRUARY 2005 VOL 307 SCIENCE www.sciencemag.org