January 31, 2006 ͉ vol. 103 ͉ no. 5 ͉ 1155–1656

Proceedings of the National Academy ofPNAS Sciences of the United States of America www.pnas.org Cover image: A bilaterally deaf, homozygous merle Shetland Sheepdog with ocular coloboma. Merle is an autosomal dominant coat pattern in the dog that is characterized by patches of diluted pigment intermingled with normal melanin. Sequence analysis reveals that this phenotype results from a retrotransposon insertion in the pigmentation gene SILV. In the homozygous state, this mutation results in a wide range of phenotypic anomalies. See the article by Clark et al. on pages 1376–1381. Image courtesy of Larry Wadsworth (Texas A&M University, College Station, TX). From the Cover

1376 Retrotransposon insertion in merle patterning 1209 Sustaining metal resources 1342 Tundra plant responses to warming 1394 Heart disease genes in Drosophila 1587 Brain histone deacetylase inhibitor

Contents PHYSICAL SCIENCES

APPLIED MATHEMATICS THIS WEEK IN PNAS 1168 Generalized multidimensional scaling: A framework for isometry-invariant partial surface matching 1155 In This Issue Alexander M. Bronstein, Michael M. Bronstein, and Ron Kimmel

COMMENTARY 1633 An auxin-driven polarized transport model for phyllotaxis 1157 Teaching an old dog new tricks: SINEs of canine Henrik Jo¨nsson, Marcus G. Heisler, Bruce E. Shapiro, genomic diversity Elliot M. Meyerowitz, and Eric Mjolsness Richard Cordaux and Mark A. Batzer ➜ See companion article on page 1376 APPLIED PHYSICAL SCIENCES 1353 Multicellularity and the functional interdependence PERSPECTIVE of motility and molecular transport Cristian A. Solari, Sujoy Ganguly, John O. Kessler, Richard E. Michod, and Raymond E. Goldstein 1159 Genetic polymorphism and protein conformational plasticity in the calmodulin superfamily: Two ways to promote multifunctionality Mitsuhiko Ikura and James B. Ames 1173 Stable and robust nanotubes stretched from polymersomes Joseph E. Reiner, Jeffrey M. Wells, Rani B. Kishore, PROFILE Candace Pfefferkorn, and Kristian Helmerson

1165 Profile of Chikashi Toyoshima 1178 Autonomous artificial nanomotor powered by sunlight ´ Regina Nuzzo Vincenzo Balzani, Miguel Clemente-Leon, Alberto Credi, ´ ➜ See Inaugural Article on page 14489 in issue 41 Belen Ferrer, Margherita Venturi, Amar H. Flood, of volume 102 and J. Fraser Stoddart

1184 Theory of the deuterium NMR of sterol– phospholipid membranes Freely available online through the PNAS open access option. Harden McConnell and Arun Radhakrishnan

PNAS ͉ January 31, 2006 ͉ vol. 103 ͉ no. 5 ͉ iii–xii Downloaded by guest on September 28, 2021 1190 Tumor marker-responsive behavior of gels prepared BIOPHYSICS by biomolecular imprinting 1173 Stable and robust polymer nanotubes stretched Takashi Miyata, Masashi Jige, Takeshi Nakaminami, from polymersomes and Tadashi Uragami Joseph E. Reiner, Jeffrey M. Wells, Rani B. Kishore, 1194 Photocatalysis and the origin of life: Synthesis Candace Pfefferkorn, and Kristian Helmerson of nucleoside bases from formamide on 1184 Theory of the deuterium NMR of sterol– TiO2(001) single surfaces S. D. Senanayake and H. Idriss phospholipid membranes Harden McConnell and Arun Radhakrishnan 1199 Cyclodextrin-covered organic nanotubes derived from self-assembly of dendrons and their 1244 Protein structure by mechanical triangulation supramolecular transformation Hendrik Dietz and Matthias Rief Chiyoung Park, Im Hae Lee, Sanghwa Lee, Yumi Song, Mikyo Rhue, and Chulhee Kim 1248 The structure of nucleosome assembly protein 1 Young-Jun Park and Karolin Luger 1215 Networks of gold nanoparticles and bacteriophage as biological sensors and cell-targeting agents 1254 Water and ligand entry in myoglobin: Assessing Glauco R. Souza, Dawn R. Christianson, Fernanda I. the speed and extent of heme pocket hydration Staquicini, Michael G. Ozawa, Evan Y. Snyder, after CO photodissociation Richard L. Sidman, J. Houston Miller, Robert A. Goldbeck, Shyam Bhaskaran, Cheri Ortega, Wadih Arap, and Renata Pasqualini Juan L. Mendoza, John S. Olson, Jayashree Soman, David S. Kliger, and Raymond M. Esquerra COMPUTER SCIENCES 1260 Torque–speed relationship of the bacterial 1627 Computer simulations reveal properties of the cell–cell flagellar motor signaling network at the shoot apex in Arabidopsis Jianhua Xing, Fan Bai, Richard Berry, and George Oster Pierre Barbier de Reuille, Isabelle Bohn-Courseau, Karin Ljung, Halima Morin, Nicola Carraro, 1266 Crystal structure of a core spliceosomal protein interface Christophe Godin, and Jan Traas Matthew J. Schellenberg, Ross A. Edwards, Dustin B. Ritchie, Oliver A. Kent, Monika M. Golas, Holger PHYSICS Stark, Reinhard Lu¨hrmann, J. N. Mark Glover, and Andrew M. MacMillan 1204 Carbon under extreme conditions: Phase boundaries and electronic properties from first-principles theory 1272 Intrinsic backbone preferences are fully present Alfredo A. Correa, Stanimir A. Bonev, and Giulia Galli in blocked amino acids Franc Avbelj, Simona Golic Grdadolnik, Joze Grdadolnik, SUSTAINABILITY SCIENCE and Robert L. Baldwin 1209 Metal stocks and sustainability 1278 Reconciling the working strokes of a single head of R. B. Gordon, M. Bertram, and T. E. Graedel skeletal muscle myosin estimated from laser-trap experiments and crystal structures BIOLOGICAL SCIENCES John Sleep, Alexandre Lewalle, and David Smith

CELL BIOLOGY 1283 Mitochondrial transfer between cells can rescue 1215 Networks of gold nanoparticles and bacteriophage aerobic respiration as biological sensors and cell-targeting agents Jeffrey L. Spees, Scott D. Olson, Mandolin J. Whitney, Glauco R. Souza, Dawn R. Christianson, Fernanda I. and Darwin J. Prockop Staquicini, Michael G. Ozawa, Evan Y. Snyder, Richard L. Sidman, J. Houston Miller, 1289 Oncogenic transformation induced by the p110␤,-␥, Wadih Arap, and Renata Pasqualini and -␦ isoforms of class I phosphoinositide 3-kinase Sohye Kang, Adam Denley, Bart Vanhaesebroeck, 1221 Long-distance lateral diffusion of human Rad51 and Peter K. Vogt on double-stranded DNA Annette Grane´li, Caitlyn C. Yeykal, Ragan B. Robertson, 1295 Nitric oxide regulates endocytosis by S-nitrosylation and Eric C. Greene of dynamin 1227 The central region of HDM2 provides a second Gaofeng Wang, Nader H. Moniri, Kentaro Ozawa, binding site for p53 Jonathan S. Stamler, and Yehia Daaka Grace W. Yu, Stefan Rudiger, Dmitry Veprintsev, Stefan Freund, M. Rosario Fernandez-Fernandez, DEVELOPMENTAL BIOLOGY and Alan R. Fersht 1301 A plausible model of phyllotaxis 1233 The Kernel Energy Method: Application to a tRNA Richard S. Smith, Soazig Guyomarc’h, Therese Mandel, Lulu Huang, Lou Massa, and Jerome Karle Didier Reinhardt, Cris Kuhlemeier, and Przemyslaw Prusinkiewicz 1238 Structural insights into a protein-bound iron-molybdenum cofactor precursor 1307 Enigma, a mitochondrial protein affecting lifespan Mary C. Corbett, Yilin Hu, Aaron W. Fay, and oxidative stress response in Drosophila Markus W. Ribbe, Britt Hedman, Philippos Mourikis, Gregory D. Hurlbut, and Keith O. Hodgson and Spyros Artavanis-Tsakonas

PNAS ͉ January 31, 2006 ͉ vol. 103 ͉ no. 5 ͉ v Downloaded by guest on September 28, 2021 1313 Real-time imaging of the somite segmentation clock: GENETICS Revelation of unstable oscillators in the individual 1376 Retrotransposon insertion in SILV is responsible presomitic mesoderm cells for merle patterning of the domestic dog Yoshito Masamizu, Toshiyuki Ohtsuka, Yoshiki Takashima, Leigh Anne Clark, Jacquelyn M. Wahl, Christine A. Rees, Hiroki Nagahara, Yoshiko Takenaka, Kenichi Yoshikawa, and Keith E. Murphy Hitoshi Okamura, and Ryoichiro Kageyama ➜ See Commentary on page 1157 1319 Coupling of cell migration with neurogenesis 1382 Active digestion of sperm mitochondrial DNA in single by proneural bHLH factors living sperm revealed by optical tweezers Weihong Ge, Fei He, Kevin J. Kim, Bruno Blanchi, Yoshiki Nishimura, Tomoya Yoshinari, Kiyoshi Naruse, Volkan Coskun, Laurent Nguyen, Xiangbing Wu, Takeshi Yamada, Kazuyoshi Sumi, Hiroshi Mitani, Jing Zhao, Julian Ik-Tsen Heng, Keri Martinowich, Tetsuya Higashiyama, and Tsuneyoshi Kuroiwa Jifang Tao, Hao Wu, Diogo Castro, Magdi M. Sobeih, Gabriel Corfas, Joseph G. Gleeson, 1388 A mutation in the human phospholamban gene, deleting Michael E. Greenberg, Francois Guillemot, arginine 14, results in lethal, hereditary cardiomyopathy and Yi E. Sun Kobra Haghighi, Fotis Kolokathis, Anthony O. Gramolini, 1325 Essential role of B-Raf in ERK activation during Jason R. Waggoner, Luke Pater, Roy A. Lynch, Guo-Chang extraembryonic development Fan, Dimitris Tsiapras, Rohan R. Parekh, Gerald W. Dorn II, Gergana Galabova-Kovacs, Dana Matzen, Daniela Piazzolla, David H. MacLennan, Dimitrios Th. Kremastinos, Katrin Meissl, Tatiana Plyushch, Adele P. Chen, Alcino Silva, and Evangelia G. Kranias and Manuela Baccarini 1394 Drosophila as a model for the identification of 1331 Intercellular coupling amplifies fate segregation during genes causing adult human heart disease Caenorhabditis elegans vulval development Matthew J. Wolf, Hubert Amrein, Joseph A. Izatt, Claudiu A. Giurumescu, Paul W. Sternberg, Michael A. Choma, Mary C. Reedy, and Anand R. Asthagiri and Howard A. Rockman 1400 The methyl-CpG binding protein MBD1 is required ECOLOGY for PML-RAR␣ function 1337 Mulberry latex rich in antidiabetic sugar-mimic alkaloids Raffaella Villa, Lluis Morey, Veronica A. Raker, Marcus forces dieting on caterpillars Buschbeck, Arantxa Gutierrez, Francesca De Santis, Kotaro Konno, Hiroshi Ono, Masatoshi Nakamura, Massimo Corsaro, Florencio Varas, Daniela Bossi, Ken Tateishi, Chikara Hirayama, Yasumori Tamura, Saverio Minucci, Pier Giuseppe Pelicci, Makoto Hattori, Akio Koyama, and Katsuyuki Kohno and Luciano Di Croce

1342 Plant community responses to experimental warming 1406 A SNP in the flt-1 promoter integrates the VEGF system across the tundra biome into the p53 transcriptional network Marilyn D. Walker, C. Henrik Wahren, Robert D. Hollister, Daniel Menendez, Oliver Krysiak, Alberto Inga, Bianca Greg H. R. Henry, Lorraine E. Ahlquist, Juha M. Alatalo, Krysiak, Michael A. Resnick, and Gilbert Scho¨nfelder M. Syndonia Bret-Harte, Monika P. Calef, Terry V. Callaghan, Amy B. Carroll, Howard E. Epstein, Ingibjo¨rg S. 1412 A genome-wide analysis of CpG dinucleotides Jo´nsdo´ttir, Julia A. Klein, Borgþo´r Magnu´sson, Ulf Molau, in the human genome distinguishes two Steven F. Oberbauer, Steven P. Rewa, Clare H. Robinson, distinct classes of promoters Gaius R. Shaver, Katharine N. Suding, Catharine C. Serge Saxonov, , and Douglas L. Brutlag Thompson, Anne Tolvanen, Ørjan Totland, P. Lee Turner, Craig E. Tweedie, Patrick J. Webber, and Philip A. Wookey 1418 Linkage disequilibrium sharing and haplotype-tagged SNP portability between populations Wei Huang, Yungang He, Haifeng Wang, Ying Wang, EVOLUTION Yangfan Liu, Yi Wang, Xun Chu, Ying Wang, Liang Xu, 1347 Climate change, body size evolution, and Cope’s Rule Yayun Shen, Xiaoyan Xiong, Hui Li, Bo Wen, Ji Qian, in deep-sea ostracodes Wentao Yuan, Chenhui Zhang, Yi Wang, Hongquan Jiang, Gene Hunt and Kaustuv Roy Guoping Zhao, Zhu Chen, and Li Jin

1353 Multicellularity and the functional interdependence 1422 Nuclear localization of the Saccharomyces cerevisiae of motility and molecular transport small subunit requires Cristian A. Solari, Sujoy Ganguly, John O. Kessler, a karyopherin and a WD40 repeat protein Richard E. Michod, and Raymond E. Goldstein Zhen Zhang, Xiuxiang An, Kui Yang, Deborah L. Perlstein, 1359 Ancient polymorphism reveals unidirectional Leslie Hicks, Neil Kelleher, JoAnne Stubbe, breeding system shifts and Mingxia Huang Boris Igic, Lynn Bohs, and Joshua R. Kohn IMMUNOLOGY 1364 Genomic fossils as a snapshot of the human transcriptome 1428 Identification of target actin content and polymerization status as a mechanism of tumor resistance after Ronen Shemesh, Amit Novik, Sarit Edelheit, cytolytic T lymphocyte pressure and Rotem Sorek Soraya Abouzahr, Georges Bismuth, Catherine Gaudin, 1370 Variable molecular clocks in hominoids Oliver Caroll, Peter Van Endert, Abdelali Jalil, Jean Dausset, Navin Elango, James W. Thomas, NISC Isabelle Vergnon, Catherine Richon, Audrey Kauffmann, Comparative Sequencing Program, Je´roˆme Galon, Graca Raposo, Fathia Mami-Chouaib, and Soojin V. Yi and Salem Chouaib

PNAS ͉ January 31, 2006 ͉ vol. 103 ͉ no. 5 ͉ vii Downloaded by guest on September 28, 2021 1434 Peptide motifs of the single dominantly expressed class I 1504 A unique role of the DNA fragmentation factor molecule explain the striking MHC-determined response in maintaining genomic stability to Rous sarcoma virus in chickens Bin Yan, Huili Wang, Yuanlin Peng, Ye Hu, He Wang, Hans-Joachim Wallny, David Avila, Lawrence G. Hunt, Xiuwu Zhang, Qi Chen, Joel S. Bedford, Mark W. Timothy J. Powell, Patricia Riegert, Jan Salomonsen, Dewhirst, and Chuan-Yuan Li Karsten Skjødt, Olli Vainio, Francis Vilbois, Michael V. Wiles, and Jim Kaufman MICROBIOLOGY 1510 Dissection of a viral autoprotease elucidates a function MEDICAL SCIENCES of a cellular chaperone in proteolysis 1440 Innate antiviral response targets HIV-1 release by T. Lackner, H.-J. Thiel, and N. Tautz the induction of ubiquitin-like protein ISG15 1516 Human ␣-defensins block papillomavirus infection Atsushi Okumura, Gengshi Lu, Ian Pitha-Rowe, Christopher B. Buck, Patricia M. Day, Cynthia D. Thompson, and Paula M. Pitha Jacek Lubkowski, Wuyuan Lu, Douglas R. Lowy, 1446 ATM promotes apoptosis and suppresses tumorigenesis and John T. Schiller in response to Myc 1522 Cleavage of the papillomavirus minor capsid protein, L2, Raju V. Pusapati, Robert J. Rounbehler, SungKi Hong, at a furin consensus site is necessary for infection John T. Powers, Mingshan Yan, Kaoru Kiguchi, Rebecca M. Richards, Douglas R. Lowy, John T. Schiller, Mark J. McArthur, Paul K. Wong, and David G. Johnson and Patricia M. Day 1452 Defective cerebellar response to mitogenic Hedgehog 1528 Identification of a conserved bacterial protein secretion signaling in Down’s syndrome mice system in Vibrio cholerae using the Dictyostelium host Randall J. Roper, Laura L. Baxter, Nidhi G. Saran, Donna K. model system Klinedinst, Philip A. Beachy, and Roger H. Reeves Stefan Pukatzki, Amy T. Ma, Derek Sturtevant, Bryan 1457 Retroviral vector integration deregulates gene Krastins, David Sarracino, William C. Nelson, expression but has no consequence on the John F. Heidelberg, and John J. Mekalanos biology and function of transplanted T cells 1534 Accordion waves in Myxococcus xanthus Alessandra Recchia, Chiara Bonini, Zulma Magnani, Oleksii Sliusarenko, John Neu, David R. Zusman, Fabrizia Urbinati, Daniela Sartori, Sara Muraro, and George Oster Enrico Tagliafico, Attilio Bondanza, Maria Teresa Lupo Stanghellini, Massimo Bernardi, Alessandra Pescarollo, Fabio Ciceri, Claudio Bordignon, NEUROSCIENCE and Fulvio Mavilio 1540 An early specific cell–cell interaction occurs in the production of ␤-amyloid in cell cultures 1463 Simultaneous molecular imaging of redox reactions Nazneen N. Dewji, Debashis Mukhopadhyay, monitored by Overhauser-enhanced MRI with and S. Jonathan Singer 14N- and 15N-labeled nitroxyl radicals Hideo Utsumi, Ken-ichi Yamada, Kazuhiro Ichikawa, 1546 Evidence that Perutz’s double-␤-stranded subunit Kiyoshi Sakai, Yuichi Kinoshita, Shingo Matsumoto, structure for ␤-amyloids also applies to their and Mika Nagai channel-forming structures in membranes S. Jonathan Singer and Nazneen N. Dewji 1469 Serpina1 is a potent inhibitor of IL-8-induced hematopoietic stem cell mobilization 1551 Olig2؉ neuroepithelial motoneuron progenitors Melissa van Pel, Ronald van Os, Gerjo A. Velders, are not multipotent stem cells in vivo Henny Hagoort, Peter M. H. Heegaard, Ivan J. D. Yoh-suke Mukouyama, Benjamin Deneen, Agne`s Lindley, Roel Willemze, and Willem E. Fibbe Lukaszewicz, Bennett G. Novitch, Hynek Wichterle, Thomas M. Jessell, and David J. Anderson 1475 Cancer-specific mutations in PIK3CA are oncogenic in vivo 1557 In vivo identification of ribonucleoprotein– Andreas G. Bader, Sohye Kang, and Peter K. Vogt RNA interactions Jennifer Zielinski, Kalle Kilk, Tiina Peritz, Theresa 1480 Pten deletion leads to the expansion of a prostatic Kannanayakal, Kevin Y. Miyashiro, Emelı´a Eirı´ksdo´ttir, stem͞progenitor cell subpopulation and tumor initiation Jeanine Jochems, Uloˆ Langel, and James Eberwine Shunyou Wang, Alejandro J. Garcia, Michelle Wu, Devon A. Lawson, Owen N. Witte, and Hong Wu 1563 Neural connections of the posteromedial cortex in the macaque 1486 Inactivation of PU.1 in adult mice leads to Josef Parvizi, Gary W. Van Hoesen, Joseph Buckwalter, the development of myeloid leukemia and Antonio Damasio Donald Metcalf, Aleksandar Dakic, Sandra Mifsud, Ladina Di Rago, Li Wu, and Stephen Nutt 1569 Morphological homeostasis in cortical dendrites Alexei V. Samsonovich and Giorgio A. Ascoli 1492 Role of ␤-arrestin 1 in the metastatic progression of colorectal cancer 1575 Organization of spines on the dendrites of Purkinje cells F. Gregory Buchanan, D. Lee Gorden, Pranathi Matta, John O’Brien and Nigel Unwin Qiong Shi, Lynn M. Matrisian, and Raymond N. DuBois 1581 A multifunctional cytoprotective agent that reduces 1498 Foamy virus vector integration sites in normal neurodegeneration after ischemia human cells Zhi-Gang Jiang, X.-C. May Lu, Valery Nelson, Xiaofang Grant D. Trobridge, Daniel G. Miller, Michael A. Jacobs, Yang, Weiying Pan, Ren-wu Chen, Michael S. Lebowitz, James M. Allen, Hans-Peter Kiem, Rajinder Kaul, Bijan Almassian, Frank C. Tortella, Roscoe O. Brady, and David W. Russell and Hossein A. Ghanbari

PNAS ͉ January 31, 2006 ͉ vol. 103 ͉ no. 5 ͉ ix Downloaded by guest on September 28, 2021 1587 The benzamide MS-275 is a potent, long-lasting brain region-selective inhibitor of histone deacetylases SOCIAL SCIENCES M. V. Simonini, L. M. Camargo, E. Dong, E. Maloku, M. Veldic, E. Costa, and A. Guidotti ANTHROPOLOGY 1593 Leptin: A potential novel antidepressant Xin-Yun Lu, Chung Sub Kim, Alan Frazer, and Wei Zhang 1370 Variable molecular clocks in hominoids Navin Elango, James W. Thomas, NISC 1599 Emotion enhances remembrance of neutral events past Comparative Sequencing Program, Adam K. Anderson, Peter E. Wais, and John D. E. Gabrieli and Soojin V. Yi

PHYSIOLOGY ECONOMIC SCIENCES 1605 Muscle GLUT4 regulation by estrogen receptors 1650 Oil market power and United States national security ER␤ and ER␣ Roger Stern Rodrigo P. A. Barros, Ubiratan F. Machado, Margaret Warner, and Jan-Åke Gustafsson POLITICAL SCIENCES 1609 A comprehensive description of the transcriptome of the hypothalamoneurohypophyseal system 1650 Oil market power and United States national security in euhydrated and dehydrated rats Roger Stern Charles Hindmarch, Song Yao, Gemma Beighton, Julian Paton, and David Murphy PSYCHOLOGY 1615 Antagonistic regulation of ROMK by long and 1599 Emotion enhances remembrance of neutral events past kidney-specific WNK1 isoforms Adam K. Anderson, Peter E. Wais, and John D. E. Gabrieli Ahmed Lazrak, Zhen Liu, and Chou-Long Huang

PLANT BIOLOGY CORRECTIONS 1621 Armadillo-related proteins promote lateral root development in Arabidopsis INAUGURAL ARTICLE, MEDICAL SCIENCES Juliet C. Coates, Laurent Laplaze, and Jim Haseloff 1656 Regulation of RNA splicing by the methylation- 1627 Computer simulations reveal properties of the cell–cell dependent transcriptional repressor signaling network at the shoot apex in Arabidopsis methyl-CpG binding protein 2 Pierre Barbier de Reuille, Isabelle Bohn-Courseau, Juan I. Young, Eugene P. Hong, John C. Castle, Juan Karin Ljung, Halima Morin, Nicola Carraro, Crespo-Barreto, Aaron B. Bowman, Matthew F. Rose, Christophe Godin, and Jan Traas Dongcheul Kang, Ron Richman, Jason M. Johnson, 1633 An auxin-driven polarized transport model Susan Berget, and Huda Y. Zoghbi for phyllotaxis Henrik Jo¨nsson, Marcus G. Heisler, Bruce E. Shapiro, BIOCHEMISTRY Elliot M. Meyerowitz, and Eric Mjolsness 1656 Structure and expression of the murine retinoblastoma POPULATION BIOLOGY gene and characterization of its encoded protein Rene Bernards, Gregory M. Shackleford, Monica R. Gerber, 1639 Secondary sex ratios and male lifespan: Jonathan M. Horowitz, Stephen H. Friend, Manfred Schartl, Damaged or culled cohorts Emil Bogenmann, Joyce M. Rapaport, Terry McGee, Ralph Catalano and Tim Bruckner Thaddeus P. Dryja, and Robert A. Weinberg

PSYCHOLOGY xiii–xiv Author Index 1644 Cortical signatures of noun and verb production Kevin A. Shapiro, Lauren R. Moo, and Alfonso Caramazza xv Subscription Form

PNAS ͉ January 31, 2006 ͉ vol. 103 ͉ no. 5 ͉ xi Downloaded by guest on September 28, 2021