DOCSLIB.ORG

Morphogenesis of Prechordal Plate and Notochord Requires Intact Eph/Ephrin B Signaling

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text
Morphogenesis of Prechordal Plate and Notochord Requires Intact Eph/Ephrin B Signaling Developmental Biology 234, 470–482 (2001) doi:10.1006/dbio.2001.0281, available online at http://www.idealibrary.com on Morphogenesis of Prechordal Plate and Notochord Requires Intact Eph/Ephrin B Signaling Joanne Chan,*,1 John D. Mably,† Fabrizio C. Serluca,† Jau-Nian Chen,† Nathaniel B. Goldstein,* Matthew C. Thomas,* Jennifer A. Cleary,* Caroline Brennan,‡ Mark C. Fishman,† and Thomas M. Roberts* *Department of Cancer Biology, Dana-Farber Cancer Institute, and Department of Pathology, Harvard Medical School, †Cardiovascular Research Center, Massachusetts General Hospital, and Department of Medicine, Harvard Medical School, Boston, Massachusetts 02115; and ‡Department of Genetics, School of Biological Sciences, Queen Mary College, University of London, Mile End Road, London E1 4NS, United Kingdom Eph receptors and their ligands, the ephrins, mediate cell-to-cell signals implicated in the regulation of cell migration processes during development. We report the molecular cloning and tissue distribution of zebrafish transmembrane ephrins that represent all known members of the mammalian class B ephrin family. The degree of homology among predicted ephrin B sequences suggests that, similar to their mammalian counterparts, zebrafish B-ephrins can also bind promiscuously to several Eph receptors. The dynamic expression patterns for each zebrafish B-ephrin support the idea that these ligands are confined to interact with their receptors at the borders of their complementary expression domains. Zebrafish B-ephrins are expressed as early as 30% epiboly and during gastrula stages: in the germ ring, shield, prechordal plate, and notochord. Ectopic overexpression of dominant-negative soluble ephrin B constructs yields reproducible defects in the morphology of the notochord and prechordal plate by the end of gastrulation. Notably disruption of Eph/ephrin B signaling does not completely destroy structures examined, suggesting that cell fate specification is not altered. Thus abnormal morphogenesis of the prechordal plate and the notochord is likely a consequence of a cell movement defect. Our observations suggest Eph/ephrin B signaling plays an essential role in regulating cell movements during gastrulation. © 2001 Academic Press Key Words: Eph; ephrin B; zebrafish; gastrulation; prechordal plate; notochord; morphogenesis; cell movement. INTRODUCTION shown to generate a mutual exclusion effect on adjacent cells allowing for the formation of rhombomere and somite Eph receptor tyrosine kinases and their ligands, the boundaries (Mellitzer et al., 1999; Xu et al., 1995; Durbin et ephrins, are cell surface molecules known to regulate cell- al., 1998). In addition, retinal axons expressing an Eph to-cell signaling. Roles for this family in developmental receptor avoid areas of high ephrin expression in the tectum processes include axon guidance, somitogenesis, angiogen- (Cheng et al., 1995; Nakamoto et al., 1996). esis, and neural crest cell migration (reviewed in Holder and Eph receptors are divided into two major classes. In Klein, 1999). Although members of this family have also general, EphA receptors bind A-ephrins that are anchored to been implicated in cell adhesion and other attraction- the plasma membrane by a glycosyl phosphatidyl-inositol related responses (reviewed in Mellitzer et al., 2000), Eph/ (GPI) linkage, and EphB receptors can bind B-ephrins that ephrin interactions have been most closely associated with have transmembrane and short, highly conserved cytoplas- repulsion-mediated cell movements. Thus, binding and mic domains. The EphA4 receptor is the only exception activation of Eph receptors and ephrin ligands have been since it can bind B-ephrins with high affinity (Brambrilla et al., 1995; Gale et al., 1996a; reviewed in Flanagan and 1 To whom correspondence should be addressed at Department Vanderhaeghen, 1998). Membrane attachment of ephrins is of Cancer Biology, Smith-970, Dana-Farber Cancer Institute, 1 thought to promote clustering of the receptors to stimulate Jimmy Fund Way, Boston, MA 02115. Fax: 617-632-4770. E-mail: cross-phosphorylation and initiation of a signal in the [email protected]. receptor-bearing cell. Furthermore, binding of either the A- 0012-1606/01 $35.00 Copyright © 2001 by Academic Press 470 All rights of reproduction in any form reserved. Eph/Ephrin B Signaling in Morphogenesis 471 or the B-ephrins to their receptors also induces signal RNA in Situ Hybridization propagation in the ligand-bearing cell (Davy and Robbins, Embryos were staged according to Kimmel et al. (1995). After 2000; Davy et al., 1999; Holland et al., 1996; Bruckner et al., fixation overnight in 4% paraformaldehyde in PBS, embryos were 1997). Thus, interaction of Ephs and ephrins leads to transferred and stored in methanol at Ϫ20°C. Whole-mount in situ bidirectional signaling with each component acting as both hybridization using digoxigenin-labeled antisense RNA probes was ligand and receptor. Soluble ephrins can act as receptor performed essentially as described by Xu et al. (1994). RNA probes antagonists since they can bind Eph receptors but do not containing digoxigenin-11-UTP and/or fluorescein-12-UTP (both induce receptor activation (Davis et al., 1994; Winslow et from Boehringer Mannheim) were synthesized from linearized al., 1995). In addition, soluble ephrins also block signal plasmid DNA. For two-color in situ labeling, both the digoxigenin- initiation in the ligand-bearing cell by occupying Eph recep- labeled and fluorescein-labeled RNA probes were added together in tors, thus, inhibiting both components of Eph/ephrin sig- the hybridization step. First, the digoxigenin probe was visualized naling. using the BM Purple alkaline phosphatase substrate (Boehringer Mannheim) to give a blue/purple color. Subsequently, the fluores- As the Eph/ephrin system has been implicated in activat- ceinated probe was detected with antifluorescein antibody (Boeh- ing cytoskeletal changes and regulating cell movement ringer Mannheim) and developed in the presence of BCIP/INT events in a number of developing systems (reviewed in Kalo (Boehringer Mannheim), for a red/orange stain. Embryos were and Pasquale, 1999; Holder and Klein, 1999), we were cleared and flat-mounted in 70% glycerol for photography. curious to see if Eph/ephrin B signaling might play an earlier role in mediating morphogenetic movements during gastrulation. Here we report the cloning and expression JB-4 Plastic Sections patterns of four members of the zebrafish ephrin B ligand Zebrafish whole-mount in situ stained embryos were dehydrated family. Each ephrin B ligand displays a dynamic expression in ethanol and infiltrated in JB-4 infiltration resin (JB-4 embedding pattern from as early as 30% epiboly to later development. solution A containing 1g/100 ml JB-4 catalyst; Poly-sciences Inc.). To examine the functional role of Eph/ephrin B signaling, After embedding the specimen in JB-4 infiltration solution [JB-4 we perturbed their interaction by microinjection of RNA Hardener (25:1) and hardening of blocks overnight at room tem- ␮ encoding dominant-negative soluble forms of B-ephrins perature], specimens were sectioned at 5 m using a Jung Supercut into the zebrafish embryo. Disruption of Eph/ephrin B 2065 microtome. signaling generated apparent morphological defects in the notochord and prechordal plate consistent with incorrect Microinjection of mRNA convergent extension movements. Using several tissue- Zebrafish ephrin B cDNAs encoding the extracellular domains specific in situ markers, we did not observe any changes in were subcloned into the pCS2ϩ vector or the pCSMT vector cell fate specification by the end of gastrulation, at the (Turner and Weintraub, 1994) and capped RNAs were in vitro tailbud stage. Our data support a role for Eph/ephrin Bs in transcribed using the Message Machine Kit (Ambion, Inc.), follow- cell movement regulation during gastrulation. ing the manufacturer’s instructions. A volume of 0.2 nl of mRNA at 100 ng/␮l encoding soluble ephrin B2b (sol-ephrin-B2b), or a frame-shifted version of soluble ephrin B2b (shift-ephrin-B2b), or a myc-tagged version of the soluble ephrin B2b (sol-ephrin-B2b-myc), MATERIALS AND METHODS or the control nuclear GFP (GFP) was microinjected into one-cell stage zebrafish embryos using a gas driven microinjector (Medical Maintenance of Zebrafish Systems Corp.). Breeding fish were maintained at 28.5°C on a 14-h light/10-h dark cycle. Embryos were collected by natural spawning, raised in RESULTS 10% Hank’s saline, and staged up to 24 h (30 somites) according to Kimmel et al. (1995); beyond this point, embryo stage is given as Sequence Analysis of Predicted Zebrafish hours postfertilization. Ephrin B Proteins Using mouse ephrin B1 and ephrin B2 cDNAs as probes, Isolation and Characterization of cDNA Clones we screened a zebrafish 18-h cDNA library at low strin- gency and isolated full-length cDNA clones encoding four The C-terminal cytoplasmic regions of mouse ephrin B1 and members of the ephrin B ligand family (Fig. 1A). Sequence ephrin B2 cDNAs (generously provided by John Flanagan) were analysis of these predicted proteins revealed structural used as probes at low stringency to screen an 18-h zebrafish homology to mammalian ephrin B1, B2, and B3; therefore, embryonic cDNA library. After three rounds of screening, four they have been named according to the degree of sequence independent full-length cDNA clones were isolated encoding four predicted proteins of the zebrafish ephrin B family: ephrin B1, homology (as assigned by the Eph nomenclature
Load more

Recommended publications
  • Late Effects of Retinoic Acid on Neural Crest and Aspects of Rhombomere Identity
    Development 122, 783-793 (1996) 783 Printed in Great Britain © The Company of Biologists Limited 1996 DEV2020 Late effects of retinoic acid on neural crest and aspects of rhombomere identity Emily Gale1, Victoria Prince2, Andrew Lumsden3, Jon Clarke4, Nigel Holder1 and Malcolm Maden1 1Developmental Biology Research Centre, The Randall Institute, King’s College London, 26-29 Drury Lane, London WC2B 5RL, UK 2Department of Molecular Biology, Princeton University, Washington Road, Princeton, NJ 08544, USA 3MRC Brain Development Programme, Division of Anatomy and Cell Biology, UMDS, Guy’s Hospital, London SE1 9RT, UK 4Department of Anatomy and Developmental Biology, University College, Windeyer Building, Cleveland St., London W1P 6DB, UK SUMMARY We exposed st.10 chicks to retinoic acid (RA), both ment of the facial ganglion in addition to a mis-direction of globally, and locally to individual rhombomeres, to look at the extensions of its distal axons and a dramatic decrease its role in specification of various aspects of hindbrain in the number of contralateral vestibuloacoustic neurons derived morphology. Previous studies have looked at RA normally seen in r4. Only this r4-specific neuronal type is exposure at earlier stages, during axial specification. Stage affected in r4; the motor neuron projections seem normal 10 is the time of morphological segmentation of the in experimental animals. The specificity of this result, hindbrain and is just prior to neural crest migration. combined with the loss of Hoxb-1 expression in r4 and the Rhombomere 4 localised RA injections result in specific work by Krumlauf and co-workers showing gain of con- alterations of pathways some crest cells that normally tralateral neurons co-localised with ectopic Hoxb-1 migrate to sites of differentiation of neurogenic derivatives.
    [Show full text]
  • Clonal Dispersion During Neural Tube Formation 4097 of Neuromeres
    Development 126, 4095-4106 (1999) 4095 Printed in Great Britain © The Company of Biologists Limited 1999 DEV2458 Successive patterns of clonal cell dispersion in relation to neuromeric subdivision in the mouse neuroepithelium Luc Mathis1,*, Johan Sieur1, Octavian Voiculescu2, Patrick Charnay2 and Jean-François Nicolas1,‡ 1Unité de Biologie moléculaire du Développement, Institut Pasteur, 25, rue du Docteur Roux, 75724 Paris Cedex 15, France 2Unité INSERM 368, Ecole Normale Supérieure, 46 rue d’Ulm, 75230 Paris Cedex 05, France *Present address: Beckman Institute (139-74), California Institute of Technology, Pasadena, CA, 91125, USA ‡Author for correspondence (e-mail: [email protected]) Accepted 5 July; published on WWW 23 August 1999 SUMMARY We made use of the laacz procedure of single-cell labelling the AP and DV axis of the neural tube. A similar sequence to visualize clones labelled before neuromere formation, in of AP cell dispersion followed by an arrest of AP cell 12.5-day mouse embryos. This allowed us to deduce two dispersion, a preferential DV cell dispersion and then by a successive phases of cell dispersion in the formation of the coherent neuroepithelial growth, is also observed in the rhombencephalon: an initial anterior-posterior (AP) cell spinal cord and mesencephalon. This demonstrates that a dispersion, followed by an asymmetrical dorsoventral (DV) similar cascade of cell events occurs in these different cell distribution during which AP cell dispersion occurs in domains of the CNS. In the prosencephalon, differences in territories smaller than one rhombomere. We conclude that spatial constraints may explain the variability in the the general arrest of AP cell dispersion precedes the onset orientation of cell clusters.
    [Show full text]
  • Mutations Affecting Cell Fates and Cellular Rearrangements During Gastrulation in Zebrafish
    Development 123, 67-80 67 Printed in Great Britain © The Company of Biologists Limited 1996 DEV3335 Mutations affecting cell fates and cellular rearrangements during gastrulation in zebrafish Lilianna Solnica-Krezel†, Derek L. Stemple, Eliza Mountcastle-Shah, Zehava Rangini‡, Stephan C. F. Neuhauss, Jarema Malicki, Alexander F. Schier§, Didier Y. R. Stainier¶, Fried Zwartkruis**, Salim Abdelilah and Wolfgang Driever* Cardiovascular Research Center, Massachusetts General Hospital and Harvard Medical School, 13th Street, Bldg. 149, Charlestown, MA 02129, USA †Present address: Department of Molecular Biology, Vanderbilt University, Box 1820, Station B, Nashville, TN 37235, USA ‡Present address: Department of Oncology, Sharett Institute, Hadassah Hospital, Jerusalem 91120, Israel §Present address: Skirball Institute of Biomolecular Medicine, NYU Medical Center, 550 First Avenue, New York, NY 10016, USA ¶Present address: School of Medicine, Department of Biochemistry and Biophysics, UCSF, San Francisco, CA 94143-0554, USA **Present address: Laboratory for Physiological Chemistry, Utrecht University, Universiteitsweg 100, 3584 CG Utrecht, The Netherlands *Author for correspondence (e-mail: [email protected]) SUMMARY One of the major challenges of developmental biology is mutations in these two groups identify genes necessary for understanding the inductive and morphogenetic processes the formation, maintenance or function of the dorsal that shape the vertebrate embryo. In a large-scale genetic organizer and the ventral signaling pathway, respectively. screen for zygotic effect, embryonic lethal mutations in Mutations in the third group affect primarily cellular zebrafish we have identified 25 mutations that affect spec- rearrangements during gastrulation and have complex ification of cell fates and/or cellular rearrangements during effects on cell fates in the embryo.
    [Show full text]
  • Stages of Embryonic Development of the Zebrafish
    DEVELOPMENTAL DYNAMICS 2032553’10 (1995) Stages of Embryonic Development of the Zebrafish CHARLES B. KIMMEL, WILLIAM W. BALLARD, SETH R. KIMMEL, BONNIE ULLMANN, AND THOMAS F. SCHILLING Institute of Neuroscience, University of Oregon, Eugene, Oregon 97403-1254 (C.B.K., S.R.K., B.U., T.F.S.); Department of Biology, Dartmouth College, Hanover, NH 03755 (W.W.B.) ABSTRACT We describe a series of stages for Segmentation Period (10-24 h) 274 development of the embryo of the zebrafish, Danio (Brachydanio) rerio. We define seven broad peri- Pharyngula Period (24-48 h) 285 ods of embryogenesis-the zygote, cleavage, blas- Hatching Period (48-72 h) 298 tula, gastrula, segmentation, pharyngula, and hatching periods. These divisions highlight the Early Larval Period 303 changing spectrum of major developmental pro- Acknowledgments 303 cesses that occur during the first 3 days after fer- tilization, and we review some of what is known Glossary 303 about morphogenesis and other significant events that occur during each of the periods. Stages sub- References 309 divide the periods. Stages are named, not num- INTRODUCTION bered as in most other series, providing for flexi- A staging series is a tool that provides accuracy in bility and continued evolution of the staging series developmental studies. This is because different em- as we learn more about development in this spe- bryos, even together within a single clutch, develop at cies. The stages, and their names, are based on slightly different rates. We have seen asynchrony ap- morphological features, generally readily identi- pearing in the development of zebrafish, Danio fied by examination of the live embryo with the (Brachydanio) rerio, embryos fertilized simultaneously dissecting stereomicroscope.
    [Show full text]
  • Hop Is an Unusual Homeobox Gene That Modulates Cardiac Development
    Cell, Vol. 110, 713–723, September 20, 2002, Copyright 2002 by Cell Press Hop Is an Unusual Homeobox Gene that Modulates Cardiac Development Fabian Chen,1 Hyun Kook,1 Rita Milewski,1 tion and are arranged in the genome in the order in Aaron D. Gitler,1,2 Min Min Lu,1 Jun Li,1 which they are expressed along the axis of the embryo. Ronniel Nazarian,1 Robert Schnepp,1 The “Hox code” represents an important paradigm for Kuangyu Jen,1 Christine Biben,3 Greg Runke,2 the definition of positional identity during embryogene- Joel P. Mackay,5 Jiri Novotny,3 sis in which the specific pattern of overlapping Hox gene Robert J. Schwartz,6 Richard P. Harvey,3,4 expression defines cellular identity. A plethora of non- Mary C. Mullins,2 and Jonathan A. Epstein1,2,7 clustered Hox genes have been identified that are scat- 1Department of Medicine tered throughout the genome and play additional roles 2 Department of Cell and Developmental Biology in cell fate specification in all organs and tissues of the University of Pennsylvania Health System body. Philadelphia, Pennsylvania 19104 Hox genes are defined by the presence of a 60 amino 3 The Victor Chang Cardiac Research Institute acid domain that mediates DNA binding. This domain Darlinghurst, NSW 2010 has been analyzed by NMR spectroscopy and X-ray Australia crystallography either free or in association with DNA 4 Faculties of Medicine and Life Sciences (Kissinger et al., 1990; Qian et al., 1989; Wolberger et University of New South Wales al., 1991). The 60 amino acid homeodomain is capable Kensington, NSW 2051 of adopting a fixed structure in solution composed of Australia three ␣ helices in which the second and third helices 5 School of Molecular and Microbial Biosciences form a helix-turn-helix motif.
    [Show full text]
  • And Krox-20 and on Morphological Segmentation in the Hindbrain of Mouse Embryos
    The EMBO Journal vol.10 no.10 pp.2985-2995, 1991 Effects of retinoic acid excess on expression of Hox-2.9 and Krox-20 and on morphological segmentation in the hindbrain of mouse embryos G.M.Morriss-Kay, P.Murphy1,2, R.E.Hill1 and in embryos are unknown, but in human embryonal D.R.Davidson' carcinoma cells they include the nine genes of the Hox-2 cluster (Simeone et al., 1990). Department of Human Anatomy, South Parks Road, Oxford OXI 3QX The hindbrain and the neural crest cells derived from it and 'MRC Human Genetics Unit, Western General Hospital, Crewe are of particular interest in relation to the developmental Road, Edinburgh EH4 2XU, UK functions of RA because they are abnormal in rodent 2Present address: Istituto di Istologia ed Embriologia Generale, embryos exposed to a retinoid excess during or shortly before Universita di Roma 'la Sapienza', Via A.Scarpa 14, 00161 Roma, early neurulation stages of development (Morriss, 1972; Italy Morriss and Thorogood, 1978; Webster et al., 1986). Communicated by P.Chambon Human infants exposed to a retinoid excess in utero at early developmental stages likewise show abnormalities of the Mouse embryos were exposed to maternally administered brain and of structures to which cranial neural crest cells RA on day 8.0 or day 73/4 of development, i.e. at or just contribute (Lammer et al., 1985). Retinoid-induced before the differentiation of the cranial neural plate, and abnormalities of hindbrain morphology in rodent embryos before the start of segmentation. On day 9.0, the RA- include shortening of the preotic region in relation to other treated embryos had a shorter preotic hindbrain than the head structures, so that the otocyst lies level with the first controls and clear rhombomeric segmentation was pharyngeal arch instead of the second (Morriss, 1972; absent.
    [Show full text]
  • Pig Antibodies
    Pig Antibodies gene_name sku Entry_Name Protein_Names Organism Length Identity CDX‐2 ARP31476_P050 D0V4H7_PIG Caudal type homeobox 2 (Fragment) Sus scrofa (Pig) 147 100.00% CDX‐2 ARP31476_P050 A7MAE3_PIG Caudal type homeobox transcription factor 2 (Fragment) Sus scrofa (Pig) 75 100.00% Tnnt3 ARP51286_P050 Q75NH3_PIG Troponin T fast skeletal muscle type Sus scrofa (Pig) 271 85.00% Tnnt3 ARP51286_P050 Q75NH2_PIG Troponin T fast skeletal muscle type Sus scrofa (Pig) 266 85.00% Tnnt3 ARP51286_P050 Q75NH1_PIG Troponin T fast skeletal muscle type Sus scrofa (Pig) 260 85.00% Tnnt3 ARP51286_P050 Q75NH0_PIG Troponin T fast skeletal muscle type Sus scrofa (Pig) 250 85.00% Tnnt3 ARP51286_P050 Q75NG8_PIG Troponin T fast skeletal muscle type Sus scrofa (Pig) 266 85.00% Tnnt3 ARP51286_P050 Q75NG7_PIG Troponin T fast skeletal muscle type Sus scrofa (Pig) 260 85.00% Tnnt3 ARP51286_P050 Q75NG6_PIG Troponin T fast skeletal muscle type Sus scrofa (Pig) 250 85.00% Tnnt3 ARP51286_P050 TNNT3_PIG Troponin T, fast skeletal muscle (TnTf) Sus scrofa (Pig) 271 85.00% ORF Names:PANDA_000462 EMBL EFB13877.1OrganismAiluropod High mobility group protein B2 (High mobility group protein a melanoleuca (Giant panda) ARP31939_P050 HMGB2_PIG 2) (HMG‐2) Sus scrofa (Pig) 210 100.00% Agpat5 ARP47429_P050 B8XTR3_PIG 1‐acylglycerol‐3‐phosphate O‐acyltransferase 5 Sus scrofa (Pig) 365 85.00% irf9 ARP31200_P050 Q29390_PIG Transcriptional regulator ISGF3 gamma subunit (Fragment) Sus scrofa (Pig) 57 100.00% irf9 ARP31200_P050 Q0GFA1_PIG Interferon regulatory factor 9 Sus scrofa (Pig)
    [Show full text]
  • Hox Genes Make the Connection
    Downloaded from genesdev.cshlp.org on September 26, 2021 - Published by Cold Spring Harbor Laboratory Press PERSPECTIVE Establishing neuronal circuitry: Hox genes make the connection James Briscoe1 and David G. Wilkinson2 Developmental Neurobiology, National Institute for Medical Research, Mill Hill, London, NW7 1AA, UK The vertebrate nervous system is composed of a vast meres maintain these partitions. Each rhombomere array of neuronal circuits that perceive, process, and con- adopts unique cellular and molecular properties that ap- trol responses to external and internal cues. Many of pear to underlie the spatial organization of the genera- these circuits are established during embryonic develop- tion of cranial motor nerves and neural crest cells. More- ment when axon trajectories are initially elaborated and over, the coordination of positional identity between the functional connections established between neurons and central and peripheral derivatives of the hindbrain may their targets. The assembly of these circuits requires ap- underlie the anatomical and functional registration be- propriate matching between neurons and the targets tween MNs, cranial ganglia, and the routes of neural they innervate. This is particularly apparent in the case crest migration. Cranial neural crest cells derived from of the innervation of peripheral targets by central ner- the dorsal hindbrain migrate ventral-laterally as discrete vous system neurons where the development of the two streams adjacent to r2, r4, and r6 to populate the first tissues must be coordinated to establish and maintain three branchial arches (BA1–BA3), respectively, where circuits. A striking example of this occurs during the they generate distinct skeletal and connective tissue formation of the vertebrate head.
    [Show full text]
  • Molekulare Untersuchungen Zur Musterbildung Im Einfachen Vielzeller Hydra
    Molekulare Untersuchungen zur Musterbildung im einfachen Vielzeller Hydra Dissertation zur Erlangung des Doktorgrades der Mathematisch-Naturwissenschaftlichen Fakultät der Christian-Albrechts-Universität zu Kiel vorgelegt von René Augustin Kiel 2003 Referent/in: ...........................................Prof. Dr. T. C. G. Bosch....... Korreferent/in: .......................................Prof. Dr. M. Leippe............... Tag der mündlichen Prüfung: ..............10.02.2004............................. Zum Druck genehmigt: Kiel, ................10.02.2004............................. Teile der vorliegenden Arbeit wurden bereits veröffentlicht oder zur Publikation vorbereitet: Fedders, H. and Augustin, R., Bosch, T.C.G. 2003. A Dickkopf-3 related gene is expressed in differentiating nematocytes in the basal metazoan Hydra. Dev. Genes Evol.: im Druck INHALTSVERZEICHNIS Inhaltsverzeichnis ABKÜRZUNGEN: 9 MOLEKULARE UNTERSUCHUNGEN ZUR MUSTERBILDUNG IM EINFACHEN VIELZELLER HYDRA 13 1. EINLEITUNG 13 1.1 Hydra als Modellorganismus der Entwicklungsbiologie 13 1.1.1 Systematik, Morphologie und Biologie 13 1.1.2 Die interstitielle Zelllinie und die Differenzierung der Nesselzellen 17 1.2 Muster- und Achsenbildung in Hydra 18 1.2.1 Musterbildung in Hydra nach Gierer & Meinhardt 19 1.2.2 Signalmoleküle in Hydra 21 1.2.3 Regulatorische Gene in Hydra 26 1.3 Zielsetzung der Arbeit 31 2. ERGEBNISSE 32 2.1 Identifizierung von HEADY Zielgenen 32 2.1.1 Ein Dickkopf homologes Protein in Hydra 32 2.1.2 Suche nach HEADY – Zielgenen mittels SSH 42 2.2 Charakterisierung der Vorgänge bei Regeneration und Knospung in Hydra 59 2.2.1 Hybridisierung /Vergleich von Filtern mit Klonen einer normalisierten cDNA Bank 59 2.2.2 Identifizierung von Unterschieden in der Genexpression bei Knospung und Regeneration in Hydra durch SSH und cDNA Macroarray Hybridisierung 67 3.
    [Show full text]
  • Regulation of Dorsal Fate in the Neuraxis by Wnt-1 and Wnt-3A
    Proc. Natl. Acad. Sci. USA Vol. 94, pp. 13713–13718, December 1997 Developmental Biology Regulation of dorsal fate in the neuraxis by Wnt-1 and Wnt-3a JEAN-PIERRE SAINT-JEANNET*†,XI HE‡§,HAROLD E. VARMUS‡, AND IGOR B. DAWID* *Laboratory of Molecular Genetics, National Institute of Child Health and Human Development, and ‡Varmus Laboratory, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892 Contributed by Igor B. Dawid, October 24, 1997 ABSTRACT Members of the Wnt family of signaling them Wnt-1 and Wnt-3a, are selectively expressed in the dorsal molecules are expressed differentially along the dorsal– but not in the ventral region of the developing nervous system ventral axis of the developing neural tube. Thus we asked (6, 12). (ii) Recent results in frog embryos have shown that whether Wnt factors are involved in patterning of the nervous Xwnt-3a is capable of inducing Krox-20 when coexpressed with system along this axis. We show that Wnt-1 and Wnt-3a, both the neural inducer noggin (13). Krox-20, a zinc-finger tran- of which are expressed in the dorsal portion of the neural tube, scription factor expressed in rhombomeres 3 and 5 of the could synergize with the neural inducers noggin and chordin hindbrain, also defines a population of cephalic neural crest in Xenopus animal explants to generate the most dorsal neural cells adjacent to rhombomere 5 that will contribute to the third structure, the neural crest, as determined by the expression of branchial arch (14). Although induction of Krox-20 by Xwnt-3a Krox-20, AP-2, and slug.
    [Show full text]
  • Formation of Regeneration of Rhombomere Boundaries in the Developing Chick Hindbrain
    Formation of regeneration of rhombomere boundaries in the developing chick hindbrain Article (Published Version) Guthrie, Sarah and Lumsden, Andrew (1991) Formation of regeneration of rhombomere boundaries in the developing chick hindbrain. Development, 112 (1). pp. 221-229. ISSN 0950- 1991 This version is available from Sussex Research Online: http://sro.sussex.ac.uk/id/eprint/69455/ This document is made available in accordance with publisher policies and may differ from the published version or from the version of record. If you wish to cite this item you are advised to consult the publisher’s version. Please see the URL above for details on accessing the published version. Copyright and reuse: Sussex Research Online is a digital repository of the research output of the University. Copyright and all moral rights to the version of the paper presented here belong to the individual author(s) and/or other copyright owners. To the extent reasonable and practicable, the material made available in SRO has been checked for eligibility before being made available. Copies of full text items generally can be reproduced, displayed or performed and given to third parties in any format or medium for personal research or study, educational, or not-for-profit purposes without prior permission or charge, provided that the authors, title and full bibliographic details are credited, a hyperlink and/or URL is given for the original metadata page and the content is not changed in any way. http://sro.sussex.ac.uk Development 112, 221-229 (1991) 221 Printed in Great Britain © The Company of Biologists Limited 1991 Formation and regeneration of rhombomere boundaries in the developing chick hindbrain SARAH GUTHRIE and ANDREW LUMSDEN Division of Anatomy and Cell Biology, United Medical and Dental Schools of Guy's and St Thomas's Hospitals, London SE1 9RT, UK Summary Development in the chick hindbrain is founded on a from adjacent positions or positions three rhombomeres segmented pattern.
    [Show full text]
  • PAX Genes in Childhood Oncogenesis: Developmental Biology Gone Awry?
    Oncogene (2015) 34, 2681–2689 © 2015 Macmillan Publishers Limited All rights reserved 0950-9232/15 www.nature.com/onc REVIEW PAX genes in childhood oncogenesis: developmental biology gone awry? P Mahajan1, PJ Leavey1 and RL Galindo1,2,3 Childhood solid tumors often arise from embryonal-like cells, which are distinct from the epithelial cancers observed in adults, and etiologically can be considered as ‘developmental patterning gone awry’. Paired-box (PAX) genes encode a family of evolutionarily conserved transcription factors that are important regulators of cell lineage specification, migration and tissue patterning. PAX loss-of-function mutations are well known to cause potent developmental phenotypes in animal models and underlie genetic disease in humans, whereas dysregulation and/or genetic modification of PAX genes have been shown to function as critical triggers for human tumorigenesis. Consequently, exploring PAX-related pathobiology generates insights into both normal developmental biology and key molecular mechanisms that underlie pediatric cancer, which are the topics of this review. Oncogene (2015) 34, 2681–2689; doi:10.1038/onc.2014.209; published online 21 July 2014 INTRODUCTION developmental mechanisms and PAX genes in medical (adult) The developmental mechanisms necessary to generate a fully oncology. patterned, complex organism from a nascent embryo are precise. Undifferentiated primordia undergo a vast array of cell lineage specification, migration and patterning, and differentiate into an STRUCTURAL MOTIFS DEFINE THE PAX FAMILY SUBGROUPS ensemble of interdependent connective, muscle, nervous and The mammalian PAX family of transcription factors is comprised of epithelial tissues. Dysregulation of these precise developmental nine members that function as ‘master regulators’ of organo- programs cause various diseases/disorders, including—and genesis4 (Figure 1).
    [Show full text]