Xenopus axis formation: induction of goosecoid by injected Xwnt-8 and activin mRNAs

Development ◽  
1993 ◽  
Vol 118 (2) ◽  
pp. 499-507 ◽  
Author(s):  
H. Steinbeisser ◽  
E.M. De Robertis ◽  
M. Ku ◽  
D.S. Kessler ◽  
D.A. Melton
Keyword(s):  
Uv Irradiation ◽  
High Frequency ◽  
Marginal Zone ◽  
Homeobox Gene ◽  
Dorsal Side ◽  
Axis Formation ◽  
Gene Products ◽  
Turn On ◽  
Xenopus Embryos ◽  
Spemann's Organizer

In this study, we compare the effects of three mRNAs-goosecoid, activin and Xwnt-8- that are able to induce partial or complete secondary axes when injected into Xenopus embryos. Xwnt-8 injection produces complete secondary axes including head structures whereas activin and goosecoid injection produce partial secondary axes at high frequency that lack head structures anterior to the auditory vesicle and often lack notochord. Xwnt-8 can activate goosecoid only in the deep marginal zone, i.e., in the region in which this organizer-specific homeobox gene is normally expressed on the dorsal side. Activin B mRNA, however, can turn on goosecoid in all regions of the embryo. We also tested the capacity of these gene products to restore axis formation in embryos in which the cortical rotation was blocked by UV irradiation. Whereas Xwnt-8 gives complete rescue of anterior structures, both goosecoid and activin give partial rescue. Rescued axes including hindbrain structures up to level of the auditory vesicle can be obtained at high frequency even in the absence of notochord structures. The possible functions of Wnt-like and activin-like signals and of the goosecoid homeobox gene, and their order of action in the formation of Spemann's organizer are discussed.

The origins of primitive blood in Xenopus: implications for axial patterning

Development ◽  
1999 ◽  
Vol 126 (3) ◽  
pp. 423-434 ◽  
Author(s):  
M.C. Lane ◽  
W.C. Smith
Keyword(s):  
Xenopus Laevis ◽  
Marginal Zone ◽  
Cell Stage ◽  
Axial Patterning ◽  
Xenopus Embryos ◽  
Spemann's Organizer ◽  
Dorsal Mesoderm

The marginal zone in Xenopus laevis is proposed to be patterned with dorsal mesoderm situated near the upper blastoporal lip and ventral mesoderm near the lower blastoporal lip. We determined the origins of the ventralmost mesoderm, primitive blood, and show it arises from all vegetal blastomeres at the 32-cell stage, including blastomere C1, a progenitor of Spemann's organizer. This demonstrates that cells located at the upper blastoporal lip become ventral mesoderm, not solely dorsal mesoderm as previously believed. Reassessment of extant fate maps shows dorsal mesoderm and dorsal endoderm descend from the animal region of the marginal zone, whereas ventral mesoderm descends from the vegetal region of the marginal zone, and ventral endoderm descends from cells located vegetal of the bottle cells. Thus, the orientation of the dorsal-ventral axis of the mesoderm and endoderm is rotated 90(degrees) from its current portrayal in fate maps. This reassessment leads us to propose revisions in the nomenclature of the marginal zone and the orientation of the axes in pre-gastrula Xenopus embryos.

scholarly journals Antimorphic goosecoids

Development ◽  
1998 ◽  
Vol 125 (8) ◽  
pp. 1347-1359 ◽  
Author(s):  
B. Ferreiro ◽  
M. Artinger ◽  
K. Cho ◽  
C. Niehrs
Keyword(s):  
Transcriptional Activation ◽  
Low Dose ◽  
Marginal Zone ◽  
Homeobox Gene ◽  
Axis Formation ◽  
Wild Type ◽  
Transcriptional Activation Domain ◽  
Spemann Organizer ◽  
Axial Defects ◽  
Dorsal Mesoderm

goosecoid (gsc) is a homeobox gene expressed in the Spemann organizer that has been implicated in vertebrate axis formation. Here antimorphic gscs are described. One antimorphic gsc (MTgsc) was fortuitously created by adding 5 myc epitopes to the N terminus of gsc. The other antimorph (VP16gsc) contains the transcriptional activation domain of VP16. mRNA injection of either antimorph inhibits dorsal gastrulation movements and leads to embryos with severe axial defects. They upregulate ventral gene expression in the dorsal marginal zone and inhibit dorsal mesoderm differentiation. Like the VP16 domain, the N-terminal myc tags act by converting wild-type gsc from a transcriptional repressor into an activator. However, unlike MTgsc, VP16gsc is able at low dose to uncouple head from trunk formation, indicating that different antimorphs may elicit distinct phenotypes. The experiments reveal that gsc and/or gsc-related genes function in axis formation and gastrulation. Moreover, this work warns against using myc tags indiscriminately for labeling DNA-binding proteins.

The Xenopus homeobox gene twin mediates Wnt induction of goosecoid in establishment of Spemann's organizer

Development ◽  
1997 ◽  
Vol 124 (23) ◽  
pp. 4905-4916 ◽  
Author(s):  
M.N. Laurent ◽  
I.L. Blitz ◽  
C. Hashimoto ◽  
U. Rothbacher ◽  
K.W. Cho
Keyword(s):  
Reporter Gene ◽  
Binding Sites ◽  
Homeobox Gene ◽  
Dorsal Side ◽  
Expression Cloning ◽  
Specific Gene ◽  
Dependent Manner ◽  
Spemann's Organizer ◽  
Dorsal Axis

We describe the isolation of the Xenopus homeobox gene twin (Xtwn), which was identified in an expression cloning screen for molecules with dorsalizing activities. Injection of synthetic Xtwn mRNA restores a complete dorsal axis in embryos lacking dorsal structures and induces a complete secondary dorsal axis when ectopically expressed in normal embryos. The sequence homology, expression pattern and gain-of-function phenotype of Xtwn is most similar to the previously isolated Xenopus homeobox gene siamois (Xsia) suggesting that Xtwn and Xsia comprise a new subclass of homeobox genes important in dorsal axis specification. We find that Xtwn is able to activate the Spemann organizer-specific gene goosecoid (gsc) via direct binding to a region of the gsc promoter previously shown to mediate Wnt induction. Since Xtwn expression is strongly induced in ectodermal (animal cap) cells in response to overexpression of a dorsalizing Wnt molecule, we examined the possibility that Xtwn might be a direct target of a Wnt signal transduction cascade. First, we demonstrate that purified LEF1 protein can interact, in vitro, with consensus LEF1/TCF3-binding sites found within the Xtwn promoter. Second, these binding sites were shown to be required for Wnt-mediated induction of a Xtwn reporter gene containing these sites. As LEF1/TCF3 family transcription factors have previously been shown to directly mediate Wnt signaling, these results suggest that Xtwn induction by Wnt may be direct. Finally, in UV-hyperventralized embryos, expression of endogenous Xtwn is confined to the vegetal pole and a Xtwn reporter gene is hyperinduced vegetally in a LEF1/TCF3-binding-site-dependent manner. These results suggest that cortical rotation distributes Wnt-like dorsal determinants to the dorsal side of the embryo, including the dorsal marginal zone, and that these determinants may directly establish Spemann's organizer in this region.

goosecoid and the organizer

Development ◽  
1992 ◽  
Vol 116 (Supplement) ◽  
pp. 167-171 ◽  
Author(s):  
Eddy M. De Robertis ◽  
Martin Blum ◽  
Christof Niehrs ◽  
Herbert Steinbeisser
Keyword(s):  
Homeobox Gene ◽  
Primitive Streak ◽  
Ventral Side ◽  
Dorsal Side ◽  
Homeodomain Protein ◽  
Spemann's Organizer ◽  
Dna Binding Specificity ◽  
Early Markers ◽  
Pharyngeal Endoderm ◽  
Molecular Nature

The molecular nature of Spemann's organizer phenomenon has long attracted the attention of embryologists. goosecoid is a homeobox gene with a DNA-binding specificity similar to that of Drosophila bicoid. Xenopus goosecoid is expressed on the dorsal side of the embryo before the dorsal lip is formed. Cells expressing goosecoid are fated to become pharyngeal endoderm, head mesoderm and notochord. Transplantation of goosecoid mRNA to the ventral side of Xenopus embryos by microinjection mimics the properties of Spemann's organizer, leading to the formation of twinned body axes, goosecoid is activated by dorsal inducers and not affected by ventral inducers. In the mouse, goosecoid is expressed in the anterior tip of the primitive streak. The availability of two early markers, goosecoid and Brachyury, opens the way for the comparative analysis of the vertebrate gastrula. The results suggest that the goosecoid homeodomain protein is an integral component of the biochemical pathway leading to Spemann's organizer phenomenon.

Microtubule disruption reveals that Spemann's organizer is subdivided into two domains by the vegetal alignment zone

Development ◽  
1997 ◽  
Vol 124 (4) ◽  
pp. 895-906 ◽  
Author(s):  
M.C. Lane ◽  
R. Keller
Keyword(s):  
Marginal Zone ◽  
Axis Formation ◽  
Convergent Extension ◽  
Microtubule Depolymerization ◽  
Microtubule Disruption ◽  
Spemann's Organizer ◽  
And Function ◽  
Cell Intercalation ◽  
Somitic Mesoderm ◽  
Anterior Posterior

Mediolateral cell intercalation is proposed to drive morphogenesis of the primary embryonic axis in Xenopus. Mediolateral intercalation begins in a group of cells called the vegetal alignment zone, a subpopulation of cells in Spemann's organizer, and spreads through much of the marginal zone. To understand the functions of the vegetal alignment zone during gastrulation and axis formation, we have inhibited its formation by disrupting microtubules with nocodazole in early gastrula embryos. In such embryos, mediolateral intercalation, involution and convergent extension of the marginal zone do not occur. Although cell motility continues, and the anterior notochordal and somitic mesoderm differentiate in the pre-involution marginal zone, posterior notochordal and somitic mesoderm do not differentiate. In contrast, microtubule depolymerization in midgastrula embryos, after the vegetal alignment zone has formed, does not inhibit mediolateral cell intercalation, involution and convergent extension, or differentiation of posterior notochord and somites. We conclude that microtubules are required only for orienting and polarizing at stage 101/2 the first cells that undergo mediolateral intercalation and form the vegetal alignment zone, and not for subsequent morphogenesis. These results demonstrate that microtubules are required to form the vegetal alignment zone, and that both microtubules and the vegetal alignment zone play critical roles in the inductive and morphogenetic activities of Spemann's organizer. In addition, our results suggest that Spemann's organizer contains multiple organizers, which act in succession and change their location and function during gastrulation to generate the anterior/posterior axis in Xenopus.

Cortical cytoplasm, which induces dorsal axis formation in Xenopus, is inactivated by UV irradiation of the oocyte

Development ◽  
1993 ◽  
Vol 119 (1) ◽  
pp. 277-285 ◽  
Author(s):  
T. Holowacz ◽  
R.P. Elinson
Keyword(s):  
Spatial Distribution ◽  
Uv Irradiation ◽  
Axis Formation ◽  
Prophase I ◽  
Xenopus Embryos ◽  
Secondary Axis ◽  
Vegetal Cortex ◽  
Dorsal Axis ◽  
Cell Embryo ◽  
Maternal Determinants

Localized maternal determinants control the formation of dorsal axial structures in Xenopus embryos. To examine the spatial distribution of dorsal determinants, we injected cytoplasm from various regions of the egg and 16-cell embryo into the ventral vegetal cells of a 16-cell recipient embryo. Cortical cytoplasm from the egg vegetal surface induced the formation of a secondary dorsal axis in 53% of recipients. In contrast, animal cortical, equatorial cortical and vegetal deep cytoplasm never induced secondary axis formation. We also compared the axis-inducing ability of animal versus vegetal dorsal cortical cytoplasm from 16-cell embryos. Significantly more dorsalizing activity was found in vegetal dorsal cytoplasm compared to animal dorsal cytoplasm at this stage. Previous work has shown that UV irradiation of the vegetal surface of either prophase I oocytes, or fertilized eggs, leads to the development of embryos that lack dorsal structures. Egg vegetal cortical cytoplasm was capable of restoring the dorsal axis of 16-cell recipient embryos derived from UV-irradiated oocytes or fertilized eggs. We also tested the axis inducing ability of cytoplasm obtained when UV-irradiated oocytes and eggs were treated as donors of cytoplasm. While vegetal cortical cytoplasm from UV-irradiated fertilized eggs retains its dorsalizing activity, cytoplasm obtained from eggs, UV irradiated as oocytes, does not. The egg vegetal cortex provides a suitable source for the isolation of maternal dorsal determinants. In addition, since UV irradiation of the oocyte vegetal surface destroys the dorsalizing activity of transferred cytoplasm, UV can be used to further restrict possible candidates for such determinants.

Dorsal downregulation of GSK3beta by a non-Wnt-like mechanism is an early molecular consequence of cortical rotation in early Xenopus embryos

Development ◽  
2000 ◽  
Vol 127 (4) ◽  
pp. 861-868 ◽  
Author(s):  
I. Dominguez ◽  
J.B. Green
Keyword(s):  
Mammalian Cells ◽  
Specific Activity ◽  
Glycogen Synthase ◽  
Dorsal Side ◽  
Dominant Negative ◽  
Axis Formation ◽  
Beta Catenin ◽  
Dorsoventral Patterning ◽  
Xenopus Embryos ◽  
Mode Of Regulation

Cortical rotation and concomitant dorsal translocation of cytoplasmic determinants are the earliest events known to be necessary for dorsoventral patterning in Xenopus embryos. The earliest known molecular target is beta-catenin, which is essential for dorsal development and becomes dorsally enriched shortly after cortical rotation. In mammalian cells cytoplasmic accumulation of beta-catenin follows reduction of the specific activity of glycogen synthase kinase 3-beta (GSK3beta). In Xenopus embryos, exogenous GSK3beta) suppresses dorsal development as predicted and GSK3beta dominant negative (kinase dead) mutants cause ectopic axis formation. However, endogenous GSK3beta regulation is poorly characterized. Here we demonstrate two modes of GSK3beta regulation in Xenopus. Endogenous mechanisms cause depletion of GSK3beta protein on the dorsal side of the embryo. The timing, location and magnitude of the depletion correspond to those of endogenous beta-catenin accumulation. UV and D(2)O treatments that abolish and enhance dorsal character of the embryo, respectively, correspondingly abolish and enhance GSK3beta depletion. A candidate regulator of GSK3beta, GSK3-binding protein (GBP), known to be essential for axis formation, also induces depletion of GSK3beta. Depletion of GSK3beta is a previously undescribed mode of regulation of this signal transducer. The other mode of regulation is observed in response to Wnt and dishevelled expression. Neither Wnt nor dishevelled causes depletion but instead they reduce GSK3beta-specific activity. Thus, Wnt/Dsh and GBP appear to effect two biochemically distinct modes of GSK3beta regulation.

XIPOU 2 is a potential regulator of Spemann's Organizer

Development ◽  
1997 ◽  
Vol 124 (6) ◽  
pp. 1179-1189 ◽  
Author(s):  
S.E. Witta ◽  
S.M. Sato
Keyword(s):  
Gene Expression ◽  
Wnt Pathway ◽  
Marginal Zone ◽  
Branchial Arch ◽  
Class Iii ◽  
Domain Family ◽  
Potent Inducer ◽  
Pou Domain ◽  
Spemann's Organizer ◽  
Dorsal Mesoderm

XIPOU 2, a member of the class III POU-domain family, is expressed initially at mid-blastula transition (MBT) and during gastrulation in the entire marginal zone mesoderm, including Spemann's Organizer (the Organizer). To identify potential targets of XIPOU 2, the interaction of XIPOU 2 with other genes co-expressed in the Organizer was examined by microinjecting XIPOU 2's mRNA into the lineage of cells that contributes to the Organizer, head mesenchyme and prechordal plate. XIPOU 2 suppresses the expression of a number of dorsal mesoderm-specific genes, including gsc, Xlim-1, Xotx2, noggin and chordin, but not Xnot. As a consequence of the suppression of dorsal mesoderm gene expression, bone morphogenetic factor-4 (Bmp-4), a potent inducer of ventral mesoderm, is activated in the Organizer. Gsc is a potential target of XIPOU 2. XIPOU 2 is capable of binding a class III POU protein binding site (CATTAAT) that is located within the gsc promoter, in the activin-inducible (distal) element. Furthermore, XIPOU 2 suppresses the activation of the gsc promoter by activin signaling. At the neurula and tailbud stages, dorsoanterior structures are affected: embryos displayed micropthalmia and the loss of the first branchial arch, as detected by the expression of pax-6, Xotx2 and en-2. By examining events downstream from the Wnt and chordin pathways, we determined that XIPOU 2, when overexpressed, acts specifically in the Organizer, downstream from GSK-3beta of the Wnt pathway and upstream from chordin. The interference in dorsalizing events caused by XIPOU 2 was rescued by chordin. Thus, in addition to its direct neuralizing ability, in a different context, XIPOU 2 has the potential to antagonize dorsalizing events in the Organizer.

Misexpression of chick Vg1 in the marginal zone induces primitive streak formation

Development ◽  
1997 ◽  
Vol 124 (24) ◽  
pp. 5127-5138 ◽  
Author(s):  
S.B. Shah ◽  
I. Skromne ◽  
C.R. Hume ◽  
D.S. Kessler ◽  
K.J. Lee ◽  
...  
Keyword(s):  
Marginal Zone ◽  
Ectopic Expression ◽  
Primitive Streak ◽  
Axis Formation ◽  
Mesoderm Induction ◽  
Cell Fates ◽  
Signalling Molecules ◽  
Posterior Area ◽  
Area Pellucida

In the chick embryo, the primitive streak is the first axial structure to develop. The initiation of primitive streak formation in the posterior area pellucida is influenced by the adjacent posterior marginal zone (PMZ). We show here that chick Vg1 (cVg1), a member of the TGFbeta family of signalling molecules whose homolog in Xenopus is implicated in mesoderm induction, is expressed in the PMZ of prestreak embryos. Ectopic expression of cVg1 protein in the marginal zone chick blastoderms directs the formation of a secondary primitive streak, which subsequently develops into an ectopic embryo. We have used cell marking techniques to show that cells that contribute to the ectopic primitive streak change fate, acquiring two distinct properties of primitive streak cells, defined by gene expression and cell movements. Furthermore, naive epiblast explants exposed to cVg1 protein in vitro acquire axial mesodermal properties. Together, these results show that cVg1 can mediate ectopic axis formation in the chick by inducing new cell fates and they permit the analysis of distinct events that occur during primitive streak formation.

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