Horst Grunz (Ed.)

The Vertebrate Organizer

Springer-Verlag Berlin Heidelberg GmbH

Horst Grunz (Ed.)

The Vertebrate Organizer

With 97 Figures, 23 in Color, and 20 Tables

Springer

Professor Dr. HORST GRUNZ FB9 Department of Zoophysiology University Duisburg-Essen UniversWitsstraBe 5 45117 Essen Germany

ISBN 978-3-642-05732-8

Library of Congress Cataloging-in-Publication Data

The vertebrate organizer I Horst Grunz (ed.). p. cm.

Includes bibliographical references. ISBN 978-3-642-05732-8 ISBN 978-3-662-10416-3 (eBook) DOI 10.1007/978-3-662-10416-3

1. Vertebrates--Embryology. 2. Organizer (Embryology) I. Grunz, Horst, 1983-

QL959.V46 2003 571.8'616--dc22

This work is subject to copyright. All rights are reserved, whether the whole or part of the material is concerned, specifically the rights of translation, reprinting, reuse of illustrations, recitation, broadcasting, reproduction on microfilms or in other ways, and storage in data banks. Duplication of this publication or parts thereof is only permitted under the provision of the German Copyright Law of September 9, 1965, in its current version, and permission for use must always be obtained from Springe.r-Verlag Berlin Heidelberg GmbH. Violations are liable for prosecution under the German Copyright Law.

© Springer-Verlag Berlin Heidelberg 2004 Originally published by Springer-Verlag Berlin Heidelberg New York in 2004 Softcover reprint of the hardcover lst edition 2004

The use of general descriptive names, registered names, trademarks, etc. in the publication does not imply, even in the absence of a specific statement, that such names are exempt from the relevant protective laws and regulations and therefore free for general use.

Cover design: Design & Production, Heidelberg Cover photographs: Whole mount in situ preparations of Xenopus embryos. Original data with photos in color see contribution of Abraham Fainsod and Vered Levy (page 102) Typesetting: Mitterweger & Partner GmbH, Plankstadt bei Heidelberg 39/3150WI - 5 4 3 2 1 0 - Printed on acid-free paper

Preface

Springer-Verlag approached me about editing a book about the Spemann­Mangold Organizer, to be published as an issue of a series concerning phy­siology. I already had some experience as an editor together with Michael Trendelenburg from the special issue, "Developmental Biology in Germany" [Int. J. Dev. Biol. 40(1), 1996]. Primarily, I wondered if it was reasonable to publish another book about the Spemann organizer so shortly after the appearance of the special issue "Spemann-Mangold Organizer [Eds. Eddy De Robertis and Juan Arechaga; Int. J. Dev. Biol. 40(1), 2000]. It could, how­ever, be argued that knowledge in this field is growing exponentially and that a lot of new data are available.

Since homologous zones of the Spemann-Mangold Organizer have mean­while been identified in zebrafish, chicken and mice, we decided that the "Ver­tebrate Organizer" would be a better title for the book. I invited many col­leagues to participate as contributors. Since the response was very positive, Springer has decided to publish the book as a separate volume.

Since the 1970s, a large increase in knowledge about the early development of the vertebrate embryo has been observed, which has accelerated dra­matically in the last decade. The first embryonic-inducing factor isolated, the vegetalizing factor, whose concentration dependently induces not only mesodermal but also endodermal tissues, was shown to be a homologue of activin A in 1990 (Grunz 1983; Green and Smith 1991). The mesoderm-induc­ing activity of the erythroid differentiation factor (identical to activin A) was discovered in 1989. In 1987, after the vegetalizing factor was shown to bind to heparin, the preferentially ventral-mesoderm-inducing activity of the hepar­in-binding growth factors (identical to fibroblast growth factors) was de­tected. The ventralizing activity of the bone morphogenetic proteins was dis­covered in amphibian embryos in 1991 and, in 1993, the nodal factor, belong­ing to the activin family, was discovered in chicken embryos. Nodal induces also mesoderm and endoderm. All these factors belong to the TGF-~ super­family. Since then, a large number of secreted factors (Chordin, Noggin, Xlim, Xvent, Cerberus, ADMP, DKK and others) as well as transcription factors (VegT, Smad's, Fox's and others) have been detected; (see also De Robertis and Wessely, this Vol.). Other approaches have used the isolation of tissues as well as the transplantation of cells to elucidate the mechanism of embryonic differentiation. The four animal cells isolated as a quartet from Xenopus eight­cell embryos, which include the future ectoderm and part of the mesoderm but no endoderm, have the ability to develop into muscle and notochord but not to endodermal tissues. After cortical rotation differentiation factors are

VI Preface

localized in the presumptive mesodermal (the so-called marginal zone) and endodermal regions of amphibian embryos. This shows that the mesoderm is determined very early and not induced by the endoderm. In early gastrulae after midblastula transition an exchange of factors between endoderm and mesoderm can occur. The same gene(s) can be expressed in adjacent parts of the mesoderm and endoderm (different germ layers). During gastrulation the definitive borders between endoderm and mesoderm are formed.

Dissociation of amphibian ectoderm followed by reaggregation leads to neural differentiation (Grunz and Tacke 1989, 1990). Obviously, neural differ­entiation is inhibited in the intact ectoderm and occurs only when this inhibi­tion is released. BMP and Wnt proteins, which inhibit neuralization of the ectoderm, can be bound by Cerberus, Dickkopf and probably other proteins (reviews: Dev. Growth Differ. 43:469-502, 2001; Int. J. Dev. Biol. 45(1), 2001; Int. J. Dev. Biol. 40(1), 1996; Naturwissenschaften 82:123-134, 1995; Blut 59:207-213, 1989).

Since the many genes and signaling pathways identified in vertebrates, in­cluding amphibians (mainly Xenopus), chicken and mice, show a high degree of identity with corresponding human genes, these discoveries are of general interest for both (molecular) biology and medicine. Since pluripotent cells (i.e. ectodermal cells) are easily available in the amphibian embryo and can be experimentally shifted into multiple pathways of differentiation, they are well suited for the study of basic molecular processes of differentia­tion.

In the last 5 years (molecular) developmental biology has been established as a core discipline of modern biology and medicine. From the 1950s until the 1970s, developmental biology (especially amphibian developmental biology correlated with the organizer phenomena) was considered a dead-end science. Nowadays, this research field has been restored to the main stream (see also review Grunz: Developmental Biology of amphibians in Germany Int. J. Dev. Biol. special issue: Spemann-Mangold organizer, 45:39-50).

Three main fields should mentioned: 1. Evolutionary developmental biology (EvoDevo) 2. Ecological developmental biology (EcoDevo) 3. Stem cell research and organ engineering

Using molecular genetic techniques, developmental biology could extend our knowledge to research fields formerly studied by traditional techniques. Comparative molecular studies of different species and even phyla have re­sulted in new insights into evolutionary conserved genes and mechanisms of differentiation. Excellent examples are the urbilateralia concept and the role of PAX genes in eye development in different species and phyla. Mean­while, the term evolutionary developmental biology (EvoDevo) is well estab­lished. There are many contributions in this volume which directly or indir­ectly address these topics.

Preface VII

Of central interest even in the popular media are the effects of chemical substances including pesticides on human health. Since the recent German Nitrofen scandal, this topic has been discussed among the general public as if it were a new topic. It might be that many people have forgotten the developmental human malformations caused by Contergan (Thalidomide) in the 1960s, or they did not imagine that harmful drugs could be a part of their daily food. Many people are discussing the possible risks of genetically manipulated plants and animals, although the acceptance of molecular genet­ics in forensic medicine is meanwhile very broad. Although Nitrofen was also found in chicken eggs, public knowledge about the correlation between sub­stances dispersed in the environment and negative influences on early em­bryonic development of most organisms is rather low. For a long time, even ecologists ignored the crucial influence of natural and anthropogenic environmental factors on early embryonic development of non-human spe­cies. In the meantime, this research field is defined as Ecological Developmen­tal Biology, even in textbooks (EcoDevo, first coined by Scott Gilbert), and research activities in this area are likely to increase exponentially in impor­tance in the coming years. Contaminations of the environment by hormones or hormone-like substances, factors interacting with receptors of the signaling pathways or reagents directly interacting with the DNA and RNA, especially during embryogenesis, are not only relevant for human fetal development, but also for invertebrates and non-human vertebrates as an important link in the food chain. Conversions of sex determination during embryonic and larval stages have already been observed in alligator and polar bear populations. For a better understanding of the risk of environmental factors on embryonic development, we need basic information about gene regulation and the com­plex signaling pathways during normogenesis. The data presented in this volume are fundamental to understanding environmental factors and pro­cesses involved in carcinogenesis and teratology.

Several chapters in this volume discuss the experimental programming of pluripotent cells to initiate different pathways of differentiation. Our group reported about organ (heart) rescue in amphibians 4 years ago (Grunz 1999), and Asashima's laboratory reported about culture and rescue experi­ments with kidney (see Chap. 15). Heart muscle with its typical honeycomb­like appearance, surrounded by an endothelial-lined pericardial cavity, can also be induced by recombinant bFGF at high concentrations in Xenopus ectoderm. Pluripotent cells from amphibians can be converted into derivatives of all three germ layers using appropriate inducing factors (activin, FGF, retinoic acid, etc.). These reports show that animal model systems can give substantial answers to basic mechanistic questions concerning the deter­mination and differentiation of pluripotent cells from all species including humans. Experimental model systems such as amphibians have the advantage that they can deliver pluripotent cells in nearly unlimited amounts. The use of human-derived stem cells only grown for basic research or human cloning issues is highly debatable and creates many ethical concerns. Other procedures

VIII Preface

for the production of human stem cells for clinical use are available (reviewed in Tiedemann, Asashima, Grunz, Knochel, Dev. Growth. Differ. 43:469-502, 2001).

Since Spemann and Hilde Mangold's famous organizer experiment in 1924, enormous progress has been made in this research field, especially following the introduction of molecular genetic techniques.

Totally new insights into mechanisms of evolution, analogy and homology relationships, and in molecular ecological studies, stem cell research and tissue/organ engineering restore developmental biology as a core discipline of modern biology and medicine.

We are optimistic that this volume will stimulate further activities of new young groups in this flourishing research field allover the world. We thank Anette Lindqvist, Editorial Assistant, for her hard work and patience turning 24 manuscripts into a book.

Essen, September 2003 HORST GRUNZ

Contents

EARLY STEPS LEADING TO THE FORMATION OF THE ORGANIZER

1 Maternal VegT and ~-Catenin: Patterning the Xenopus Blastula. Matthew Kofron, Jennifer Xanthos, and Janet Heasman

1.1 Introduction. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 1 1.2 Cell Fate Specification in the Animal-Vegetal Axis ......... " 1 1.2.1 Endodermal Transcription Factors Downstream of

VegT Have General and Specific Roles in Fate Specification . . .. 4 1.2.2 The Importance of Inductive Interactions in Mesoderm

and Endoderm Specification . . . . . . . . . . . . . . . . . . . . . . . . . .. 4 1.3 Patterning in the Dorso-Ventral Axis ................... " 5 1.4 Conclusions . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 8

References ...................................... " 9

2 Short-Versus Long-Range Effects of Spemann's Organizer. . . .. 11 Ira 1. Blitz and Ken W. Y. Cho

2.1 Introduction. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 11 2.2 What Are the Organizer-Derived Dorsalizing Signal(s)? . . . . . .. 13 2.3 Are the Long-Range Effects of the Organizer Really Long Range? 15 2.3.1 The Heart Primordia and Anterior Somites Are Specified

by Short-Range Signaling During Gastrulation. . . . . . . . . . . . .. 16 2.3.2 Specification of the Pronephros Provides an Example

of a Secondary Induction Occurring During Late Gastrulation .. 17 2.3.3 Specification of Posterior Somites Provides an Example

of Late Short-Range Induction by Organizer-Derived Structures. 18 2.4 Concluding Remarks. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 19

References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 20

3 Formation of the Endoderm in Xenopus. . . . . . . . . . . . . . . . .. 25 Hugh R. Woodland and Debbie Clements

3.1 Intro duction. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 25 3.1.1 Concepts and Views of Patterning of the Embryo. . . . . . . . . . .. 25 3.2 Early Endoderm Formation in Xenopus. . . . . . . . . . . . . . . . . .. 27 3.2.1 Phases of Endoderm Formation. . . . . . . . . . . . . . . . . . . . . . . .. 27 3.2.2 The Initiation/Maternal Phase of VegT Action .. . . . . . . . . . . .. 29 3.2.3 The Establishment of the Endoderm . . . . . . . . . . . . . . . . . . . .. 29 3.2.4 Why Are There So Many Signalling Molecules Involved

in the Endoderm Community Effect? . . . . . . . . . . . . . . . . . . . .. 31

X Contents

3.2.5 The Role of VegT Targets in Endoderm Formation. . . . . . . . . .. 32 3.2.6 Delimitation of the Endodermal Domain. . . . . . . . . . . . . . . . .. 35 3.3 Patterning of the Endoderm . . . . . . . . . . . . . . . . . . . . . . . . . .. 35 3.4 Conclusions . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 36

References. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 37

4

4.1 4.2 4.2.1 4.2.2 4.2.3 4.3 4.3.1 4.3.1.1 4.3.1.2 4.3.1.3 4.3.2 4.3.2.1 4.3.3 4.3.3.1 4.4

Role of Fox Genes During Xenopus Embryogenesis . ......... 41 Hsiu-Ting Tseng, Isaac Brownell, Ryuju Hashimoto, Heithem El-Hodiri, Olga Medina-Martinez, Rina Shah, Carolyn Zilinski, and Milan Jamrich

Introduction. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 41 Expression of Fox Genes in the Mesoderm. . . . . . . . . . . . . . . .. 42 FoxA4 (XFKHl) .................................... 44 FoxC2 (XFKH7) . .................................... 44 FoxFl (XFD-13) .................................... 45 Expression of Fox Genes in the Ectoderm . . . . . . . . . . . . . . . .. 46 Neuroectoderm ..................................... 46 FoxGl (XFKH4/ XBF-l) . .............................. 47 FoxB2 (XFD-S) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 48 FoxD3 (XFD6/ XFKH6) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 48 Placodal Ectoderm . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 49 FoxE4 (Xlensl) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 49 Epidermis. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 50 Fox!l (XFKHS) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 50 Conclusions . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 51 References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 51

THE ROLE OF THE ORGANIZER

5 The Molecular Nature of Spemann's Organizer. . . . . . . . . . . .. 55 E. M. De Robertis and Oliver Wessely

5.1 Historical Background. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 55 5.2 The Spemann Organizer Transcription Factors. . . . . . . . . . . . .. 55 5.3 The Organizer Secreted Factors. . . . . . . . . . . . . . . . . . . . . . . .. 57 5.3.1 TGFp Superfamily Antagonists. . . . . . . . . . . . . . . . . . . . . . . . .. 58 5.3.1.1 The Chordin and Noggin BMP Antagonists ................. 58 5.3.1.2 Gremlin and Sclerostin . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 58 5.3.1.3 Follistatin......................................... 59 5.3.1.4 Xnr-3............................................ 59 5.3.1.5 Lefty and Antivin . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 59 5.3.1.6 Cerberus......................................... 60 5.3.1.7 Secreted Wnt Inhibitors . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 61 5.4 Chordin and the Organizer . . . . . . . . . . . . . . . . . . . . . . . . . . .. 63 5.5 The Chordin Co-factors. . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 65 5.6 Neural Induction and the Spemann Organizer. . . . . . . . . . . . .. 66

References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 68

Contents

6

6.1 6.2 6.2.1 6.2.2 6.2.2.1

6.2.2.2 6.2.2.3

6.2.2.4

6.2.3 6.2.3.1

6.2.3.2

6.3 6.4 6.5 6.6 6.7

7

7.1 7.2

7.2.1 7.2.2 7.2.3 7.3 7.3.1 7.3.2 7.3.3 7.4 7.4.1 7.4.2 7.4.3 7.5

XI

The Community Effect in Xenopus Development. . . . . . . . . . .. 73 Henrietta J. Standley and J. B. Gurdon

Introduction. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 73 The Community Effect in Xenopus Myogenesis. . . . . . . . . . . . .. 75 A Community Effect Is Required for Muscle Differentiation . . .. 75 The Search for the Myogenic Community Factor . . . . . . . . . . .. 76 The Myogenic Community Factor May Be Secreted or Associated with the Cell Membrane. . . . . . . . . . . . . . . . . . .. 76 Organiser Factors Cannot Account for the Community Effect. .. 77 Candidate Community Factors Can Be Tested for Community Activity by Supplying Them in Protein Form to Dispersed Cells. 78 eFGF Fulfills the Criteria Demanded of the Endogenous Myogenic Community Factor. . . . . . . . . . . . . . . . . . . . . . . . . .. 80 The Timing of the Community Effect. . . . . . . . . . . . . . . . . . . .. 81 The Community Effect is Operative During Gastrulation and Neurulation .................................... 81 The Period of Sensitivity to eFGF Coincides with the Endogenous Community Interaction. . . . . . . . . . . . . . . . . . . .. 81 The Community Effect in the Notochord. . . . . . . . . . . . . . . . .. 82 The Community Effect and Mesoderm Maintenance. . . . . . . . .. 83 The Mechanism of the Community Effect. . . . . . . . . . . . . . . . .. 84 Future Directions . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 88 Concluding Remarks. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 89 References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 89

Regulation of Spemann's Organizer Formation. . . . . . . . . . . .. 93 Abraham Fainsod and Vered Levy

Introduction. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 93 Positive Regulation of Organizer Formation -Establishing Spemann's Organizer . . . . . . . . . . . . . . . . . . . . . .. 94 The Wnt Signaling Pathway. . . . . . . . . . . . . . . . . . . . . . . . . . .. 94 Positioning the Wnt Signal for Organizer Induction. . . . . . . . .. 95 Timing of the Early Wnt Signal. . . . . . . . . . . . . . . . . . . . . . . .. 97 BMP Signaling as a Negative Regulator of Organizer Formation. 98 Inhibition of BMP Signaling Induces Secondary Axes. . . . . . . .. 98 Localization and Timing of the Early BMP Signal. . . . . . . . . . .. 99 BMP Downstream Effectors. . . . . . . . . . . . . . . . . . . . . . . . . .. 100 Competence of the Marginal Zone To Become Organizer Tissue 101 Timing of Secondary Axis Induction. . . . . . . . . . . . . . . . . . .. 101 Controlling the Ventral Marginal Zone Competence. . . . . . . .. 104 Temporal and Spatial Expression of Xcad2 . . . . . . . . . . . . . . .. 106 Establishment of the Organizer - a Model. . . . . . . . . . . . . . .. 107 References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 107

XII Contents

8 Transcriptional Repression in Spemann's Organizer and the Formation of Dorsal Mesoderm. . . . . . . . . . . . . . . .. 113 Sergey Yaklichkin, Aaron B. Steiner, and Daniel S. Kessler

8.1 Introduction. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 113 8.1.1 Xenopus Mesoderm Formation: Embryological Analysis. . . . .. 113 8.1.2 Xenopus Mesoderm Formation: Molecular Analysis ......... 114 8.1.3 The Fox Gene Family. . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 115 8.1.4 FoxD3 in the Vertebrate Embryo. . . . . . . . . . . . . . . . . . . . . .. 115 8.2 Results. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 116 8.2.1 Embryonic Expression of Xenopus FoxD3. . . . . . . . . . . . . . . .. 116 8.2.2 Regulation of FoxD3 by the Nodal and Wnt Signaling Pathways 118 8.2.3 Dorsal Mesoderm Induction by FoxD3. . . . . . . . . . . . . . . . . .. 120 8.2.4 Nuclear Localization and Transcriptional Activity of FoxD3 . .. 121 8.3 Conclusions. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 123

References. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 124

9 Wnt Signals and Antagonists: The Molecular Nature of Spemann's Head Organizer . . . . . .. 127 Christof Niehrs

9.1 Summary. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 127 9.2 The Spemann Organizer. . . . . . . . . . . . . . . . . . . . . . . . . . . .. 127 9.3 Head, Trunk and Tail Organizers Versus Gradient Models. . . .. 128 9.4 Wnt/~-Catenin Signalling Antagonizes

Spemann's Head Organizer . . . . . . . . . . . . . . . . . . . . . . . . . .. 130 9.5 Production of Wnt Antagonists is a Distinguishing Function

of the Head Organizer. . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 132 9.6 Wnt Antagonists and Patterning of Mesoderm and Endoderm.. 135 9.7 A Posteriorizing Gradient of Wnt/~-Catenin Activity

Regulates A-P Patterning. . . . . . . . . . . . . . . . . . . . . . . . . . . .. 136 9.8 Interactions Between Wnt, BMP, FGF and RA Signalling

in A-P Patterning. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 139 9.8.1 BMPs . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 139 9.8.2 FGFs. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 140 9.8.3 Retinoic Acid (RA) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 140 9.9 Orthogonal Signalling Gradients of Wnt and BMP

Specify A-P and D-V Embryonic Axes. . . . . . . . . . . . . . . . . .. 141 9.10 Conclusions. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 142

References. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 143

Contents XIII

10 Basic Helix-Loop-Helix Proneural Genes and Neurogenesis in Xenopus Embryos. . . . . . . . . . . . . . . . .. 151 Eric Bellefroid and Jacob Souopgui

10.1 Introduction. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 151 10.2 Proneural Genes in Xenopus. . . . . . . . . . . . . . . . . . . . . . . . .. 154 10.3 Transcriptional Factors Controlling the Activation

of bHLH Proneural Genes. . . . . . . . . . . . . . . . . . . . . . . . . . .. 155 10.4 Proneural Genes and the Selection of Neuronal Progenitors . .. 156 10.5 Cascade of Proneural-Differentiation Genes. . . . . . . . . . . . . .. 160 10.6 Stabilization of the Neuronal Differentiation Process . . . . . . .. 160 10.7 Downstream Targets of Proneural Factors

Regulating Later Steps in Neuronal Differentiation. . . . . . . . .. 162 10.8 Proneural Factors Promote Cell Cycle Exit. . . . . . . . . . . . . . .. 163 10.9 Inhibition of Glia/Neural Crest Fate and Neuronal

Subtype Specification. . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 165 10.10 Conclusions and Prospects. . . . . . . . . . . . . . . . . . . . . . . . . .. 167

References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 168

11 Organizer Activities Mediated by Retinoic Acid Signaling. . .. 173 Yonglong Chen, Thomas Hollemann, and Tomas Pieler

11.1 Summary . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 173 11.2 Introduction. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 173 11.3 RA Generation and Degradation . . . . . . . . . . . . . . . . . . . . . .. 175 11.4 RALDH2 and CYP26A1 May Generate a Dynamic RA

Concentration Gradient In Vivo. . . . . . . . . . . . . . . . . . . . . . .. 177 11.5 RA Signaling and Hindbrain Patterning. . . . . . . . . . . . . . . . .. 179 11.6 Concluding Remarks. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 183

References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 183

12 Wnt Signalling and Regulation of Gastrulation Movements.. 187 De-Li Shi

12.1 12.1.1 12.1.2 12.2 12.2.1 12.2.2

12.2.3

12.2.4 12.2.5

Introduction. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 187 Convergent Extension Movements in Gastrulation . . . . . . . . .. 187 Wnt Signalling Pathways. . . . . . . . . . . . . . . . . . . . . . . . . . . .. 189 Molecular Regulation of Convergent Extension Movements. . .. 190 Wnt Signalling Activity and Convergent Extension Movements. 190 The JNK Pathway Regulates Convergent Extension During Gastrulation. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 191 Wnt Signalling in Neural Convergent Extension and Neural Fold Closure. . . . . . . . . . . . . . . . . . . . . . . . . . . .. 194 The Wnt/Ca2+ Pathway Regulates Cell-Cell Adhesion. . . . . . .. 194 Otx-2 and Inhibition of Convergent Extension in Head Mesoderm. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 195

12.3 Conclusions. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 196 References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 196

XIV Contents

THE ORGANIZER AND PATTERN FORMATION

13 How Cell-Cell Adhesion Contributes to Early Embryonic Development. . . . . . . . . . . . . . . . . . . . .. 201 Gui Ruan, Doris Wedlich, and Almut Kohler

13.1 Introduction ...................................... 201 13.2 Who Is Who Among Xenopus Cadherins? . . . . . . . . . . . . . . .. 203 13.2.1 XB/U-Cadherin and EP/C-Cadherin. . . . . . . . . . . . . . . . . . . .. 203 13.2.2 Epithelial Cadherins . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 204 13.2.3 Neural Cadherins . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 205 13.2.4 Type II Cadherins. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 205 13.2.5 Protocadherins.................................... 206 13.3 Functions of Cadherins in Xenopus. . . . . . . . . . . . . . . . . . . .. 206 13.3.1 Cadherin-Catenin Complexes During Cleavage

and Blastula Stage. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 207 13.3.2 Cadherins Control Cell Movements During Gastrulation. . . . .. 208 13.3.3 How Is Adhesive Strength Modulated in Cell Movements? . . . .. 210 13.3.4 Cadherins in Tissue Formation. . . . . . . . . . . . . . . . . . . . . . .. 211 13.4 Perspectives. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 214

References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 215

14 Patterning Non-neural Ectoderm by Organizer-Modulated Homeodomain Factors. . . . . . . . . .. 219 Thomas D. Sargent

14.1 Introduction. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 219

14.2 Axial Specificity of Msx and Dlx Gene Expression in the Ectoderm 221 14.3 Msx and Dlx Genes: Local Indicators of BMP Signal Strength.. 223 14.4 Selective Repression of Ectodermal Targets by Msx/Dlx Genes. 225 14.5 A Homeobox Gene Code for Ectodermal Patterning. . . . . . . .. 226

References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 230

AXIS FORMATION AND ORGANOGENESIS

15 Embryonic Organogenesis and Body Formation in Amphibian Development . . . . . . . . . . . . . . . . . . . . . . . . .. 233 M. Asashima, A. Sogame, T. Ariizumi, and T. Igarashi

15.1 Introduction. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 233 15.2 Animal Cap Assay and Activin Findings. . . . . . . . . . . . . . . . .. 234 15.3 Formation of Heart. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 238 15.4 Formation of Kidney. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 240 15.5 Formation of Pancreas . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 243 15.6 Formation of Cartilage. . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 245 15.7 Formation of Central Nervous System and Sensory Organs:

In Vitro Control of Embryonic Axis Formation . . . . . . . . . . .. 246

Contents XV

15.8 Formation of Heads and Trunk-and-Tails: In Vitro Control of Embryonic Body Plan by Artificial Organizer 248

15.9 Advancing from the Basic to Applied Stages. . . . . . . . . . . . . .. 250 References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 252

16 Organizing the Eye. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 257 Robert Vignali, Massimiliano Andreazzoli, Federico Cremisi, and Giuseppina Barsacchi

16.1 Introduction. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 257 16.2 Neural Induction and the Initial Eye Field. . . . . . . . . . . . . . .. 258 16.3 Splitting of the Early Eye Field . . . . . . . . . . . . . . . . . . . . . . .. 261 16.4 Eye Field Patterning and the Role of Eye Field Specific

Transcription Factors . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 263 16.5 Factors Establishing Eye Polarity. . . . . . . . . . . . . . . . . . . . . .. 267 16.6 Control of Proliferation During Retinogenesis . . . . . . . . . . . .. 268 16.7 Retinal Cell Fate Specification ....................... " 269 16.8 Concluding Remarks .............................. " 273 16.9 Acknowledgements. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 274

References ..................................... " 274

17 A Critical Role for Retinoid Receptors in Axial Patterning and Neuronal Differentiation . ....... " 279 Andres E. Carrasco and Bruce Blumberg

17.1 Retinoid Signaling . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 279 17.2 Anteroposterior Neural Patterning. . . . . . . . . . . . . . . . . . . . .. 280 17.3 Retinoid Signaling in AlP Patterning. . . . . . . . . . . . . . . . . . .. 282 17.4 W nt, FGF and Retinoid Signaling Converge on Xcad3 ...... " 283 17.5 Interaction Between RAR and FGF Signaling

During Xenopus AlP Patterning ...................... " 284 17.6 RA Signaling Is Involved in Multiple Steps

in Posteriorization of the Neural Tube. . . . . . . . . . . . . . . . . .. 285 17.7 Neurogenesis - a Brief Story. . . . . . . . . . . . . . . . . . . . . . . . .. 286 17.8 RA Induces the Expression of N -Tubulin in the Neural Plate. .. 288 17.9 The RA-Mediated Increase in Primary Neurons Does Not Result

from Alterations in Proliferation or Apoptosis . . . . . . . . . . . .. 289 17.10 Retinoids Affect the Expression of Genes

Involved in Primary Neurogenesis . . . . . . . . . . . . . . . . . . . . .. 289 17.11 Shh Signaling Delays Neuronal Differentiation. . . . . . . . . . . .. 290 17.12 Retinoids Inhibit shh Expression

to Promote Neuronal Differentiation. . . . . . . . . . . . . . . . . . .. 291 17.13 Conclusions: Retinoids Regulate Early AlP Patterning

and Early Steps in the Neurogenic Cascade ............. " 292 References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 293

XVI Contents

18 Molecular Patterning of the Embryonic Brain. . . . . . . . . . . .. 299 Esther Bell and Ali H. Brivanlou

lS.l Introduction. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 299 lS.2 Neural Induction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 300 lS.3 Initiation of the Neural Patterning. . . . . . . . . . . . . . . . . . . . .. 301 lS.4 Induction and Patterning of the Forebrain

along the AP, DV and LR Axes. . . . . . . . . . . . . . . . . . . . . . . .. 302 lS.4.1 Establishment of the Telencephalon and Diencephalon:

Putative Signaling Centers and Molecular Pathways . . . . . . . .. 302 lS.4.2 Regulatory Genes That Pattern the Forebrain Along the DV Axis 30S lS.4.3 Is the Forebrain a Segmented Structure? . . . . . . . . . . . . . . . .. 30S lS.4.4 Patterning of the LR Axis of the Forebrain. . . . . . . . . . . . . . .. 306 lS.S Midbrain Patterning. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 306 lS.S.l The DV Axis. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 306 lS.S.2 The AP Axis. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 307 lS.6 Hindbrain Patterning. . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 307 lS.6.1 The AP Axis. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 307 lS.6.2 The DV Axis of the Hindbrain. . . . . . . . . . . . . . . . . . . . . . . .. 30S lS.7 Conclusions. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 309

19

19.1 19.2 19.2.1 19.2.2

19.2.3

19.2.4

19.2.S

19.2.6

19.2.7 19.2.S

19.2.9 19.3

19.3.1 19.3.1.1 19.3.1.2 19.3.2

References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 309

Epidermal, Neuronal and Glial Cell Fate Choice in the Embryo 31S Marc Moreau, Philippe Cochard, and Anne-Marie Duprat

Introduction. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 31S Neural Cell Fate Determination. . . . . . . . . . . . . . . . . . . . . . .. 316 What Are the Neuralizing Factors? . . . . . . . . . . . . . . . . . . . .. 316 Dissociation Inhibits Epidermal Fate and Neuralizes the Ectodermal Cells. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 316 Calcium Is Involved in the Choice Between Neural and Epidermal Fate. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 319 Neuralization by Dissociation of Ectodermal Cells Is Associated with a Calcium Signal. . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 319 Noggin Triggers an Increase in Intracellular Calcium Concentration . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 320 Direct Activation of L-Type Calcium Channels Triggers Neural Induction on Isolated Ectoderms. . . . . . . . . .. 321 Targets of Calcium . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 323 Neural Competence of the Ectoderm Is Linked to the Expression of L-Type Calcium Channels . . . . . . . . . . . . . . . . . . . . . . . . .. 323 Control of the Choice of Determination: a New Model . . . . . .. 324 Specification Mechanisms of Glial Cells in the Central Nervous System. . . . . . . . . . . . . . . . . . . . . . . .. 326 Specification Mechanisms of Astrocytes. . . . . . . . . . . . . . . . .. 327 The Stem Cell Potential of Astrocytes . . . . . . . . . . . . . . . . . .. 327 The Switch Between Neuronal and Astroglial Fates. . . . . . . . .. 32S Mechanisms of Oligodendrocyte Specification . . . . . . . . . . . .. 330

Contents

19.3.2.1 19.3.2.2 19.3.2.3 19.4

Oligodendrocyte Origins ............................ . Oligodendrocyte Induction .......................... . Negative Control of Oligodendrocyte Specification ......... . Concluding Remarks ............................... . References ...................................... .

XVII

330 331 333 335 336

20 Downstream of the Organizer: The Xenopus Cement Gland as a Model for Organ Positioning and Differentiation . . . . .. 343 Fiona C. Wardle and HazelL. Sive

20.1 Introduction. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 343 20.2 The Cement Gland . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 343 20.3

20.3.1 20.3.2 20.4

20.4.1 20.4.2

Positioning the Cement Gland: Activity of Positive and Negative Tissue Interactions and Secreted Signals. . . . . .. 345 Positive Factors .................... . . . . . . . . . . . . . .. 346 Inhibitory Factors. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 347 Positioning the Cement Gland Requires the Overlap of Three Larger Domains ............................ 347 The Venn Diagram Model. . . . . . . . . . . . . . . . . . . . . . . . . . .. 347 AD: The Anterodorsal Domain. . . . . . . . . . . . . . . . . . . . . . . .. 348

20.4.3 VL: The Ventrolateral Domain. . . . . . . . . . . . . . . . . . . . . . . .. 350 20.4.4 EO: The Outer Ectodermal Domain. . . . . . . . . . . . . . . . . . . .. 351 20.5 Intermediate Domains in Cement Gland Formation . . . . . . . .. 351 20.5.1 VL+E: A Ventral Ectodermal Domain. . . . . . . . . . . . . . . . . . .. 352 20.5.2 AD+VL+E: An Anterior Ectodermal Domain. . . . . . . . . . . . .. 352 20.6 Differentiation of the Cement Gland: Identification of Factors

That Control Cement Gland Differentiation Genes . . . . . . . . .. 352 20.6.1 Promoter Analysis of the Cement Gland Specific Gene, Xagl. .. 353 20.6.2 Does Expression of All Cement Gland Differentiation

Genes Require the Same Transcription Factors? . . . . . . . . . . .. 353 20.7 From Position to Differentiation. . . . . . . . . . . . . . . . . . . . . .. 354

References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 354

ORGANIZER FORMATION IN DIFFERENT VERTEBRATES

21 The Organizer in Amphibians with Large Eggs:

21.1 21.2 21.3 21.3.1 21.3.2 21.4 21.5 21.5.1 21.5.2

Problems and Perspectives. . . . . . . . . . . . . . . . . . . . . . . . . .. 359 Eugenia M. del Pino and Richard P. Elinson

Introduction. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 359 Large and Small Amphibian Eggs .............. . . . . . . .. 360 Slow and Rapid Development in Frogs with Large Eggs . . . . .. 362 Slow Development in G. riobambae . . . . . . . . . . . . . . . . . . . .. 362 Rapid Development in E. coqui and Other Frogs. . . . . . . . . . .. 363 Mesoderm Formation in Frogs with Large Eggs . . . . . . . . . . .. 364 Formation of the Organizer. . . . . . . . . . . . . . . . . . . . . . . . . .. 365 The Organizer in G. riobambae . . . . . . . . . . . . . . . . . . . . . . .. 368 The Organizer in E. coqui . . . . . . . . . . . . . . . . . . . . . . . . . . .. 369

XVIII Contents

21.6 Concluding Remarks. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 371 References. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 371

22 Formation and Functions of the Gastrula Organizer in Zebrafish 375 Joshua S. Waxman and Randall T. Moon

22.1 Introduction. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 375 22.2 Early Observations and Experimental Studies

of the Teleost Gastrula Organizer. . . . . . . . . . . . . . . . . . . . . .. 375 22.3 Formation of the Teleost Organizer. . . . . . . . . . . . . . . . . . . .. 377 22.4 Physical Components Contributing to the Formation

of the Organizer. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 378 22.5 The Teleost Nieuwkoop Center, Or Not? . . . . . . . . . . . . . . . . .. 379 22.6 Genes That Function in the Teleost Gastrula Organizer. . . . . .. 381 22.6.1 Bozozok......................................... 381 22.6.2 The Nodals. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 382 22.6.3 Dickkopf........................................ 383 22.6.4 Hhex........................................... 384 22.6.5 Chordin......................................... 384 22.7 Downstream of boz and the Nodals; Partners in Specifying

the Organizer. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 385 22.8 Antagonistic Interactions among Organizer

Vs Non-organizer Signals. . . . . . . . . . . . . . . . . . . . . . . . . . .. 386 22.9 Interactions Between boz, eye and sqt . . . . . . . . . . . . . . . . . .. 387 22.10 Interactions among Ventral-Lateral Specification Genes:

wntB and BMPs. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 388 22.11 Organizer Activity and the Control

of Morphogenetic Movements . . . . . . . . . . . . . . . . . . . . . . . .. 389 22.12 Conclusions. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 389

References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 390

23 Hensen's Node: The Embryonic Organizer of the Chick. . . . .. 395 Lars Wittler, Derek Spieler, and Michael Kessel

23.1 Introduction. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 395 23.2 From the Unicellular Embryo to the Single-Layered Blastoderm 396 23.3 The Two Layers of the Avian Blastula. . . . . . . . . . . . . . . . . . .. 399 23.4 The Primitive Streak of the Early Avian Gastrula. . . . . . . . . .. 400 23.5 Hensen's Node in the Late Gastrula. . . . . . . . . . . . . . . . . . . .. 400 23.6 The Prechordal Plate/Mesendoderm of the Chick Embryo. . . .. 403 23.7 The Establishment of the Left-Right Axis. . . . . . . . . . . . . . . .. 403 23.8 Conclusions and Outlook . . . . . . . . . . . . . . . . . . . . . . . . . . .. 405

References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 406

Contents

24

24.1 24.2 24.3 24.3.1 24.3.2 24.3.3 24.4 24.4.1 24.4.2

XIX

Formation and Function of the Mouse Organizer. . . . . . . . .. 409 Luc Leyns and Caroline R. Kemp

Introduction. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 409 Early Mouse Embryonic Development . . . . . . . . . . . . . . . . . .. 409 Formation of a Double Axis in Mammals. . . . . . . . . . . . . . . .. 412 Polyembryony. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 412 Experimental Manipulations . . . . . . . . . . . . . . . . . . . . . . . . .. 413 Molecular Manipulations. . . . . . . . . . . . . . . . . . . . . . . . . . . .. 414 Molecular Inductions . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 415 Establishment of the Proximal-Distal Axis. . . . . . . . . . . . . . .. 416 Formation of the Anterior-Posterior Axis. . . . . . . . . . . . . . . .. 418 References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 420

Subject Index. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. 423