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PubMed2001

The pitx2 homeobox protein is required early for endoderm formation and nodal signaling. .

Nodal and Nodal-related factors play fundamental roles in a number of developmental processes, including mesoderm and endoderm formation, patterning of the anterior neural plate, and determination of bilateral asymmetry in vertebrates. pitx2, a paired-like homeobox gene, has been proposed to act downstream of Nodal in the gene cascade providing left-right cues to the developing organs. Here, we report that pitx2 is required early in the Nodal signaling pathway for specification of the endodermal and mesodermal germ layers. We found that pitx2 is expressed very early during Xenopus and zebrafish development and in many regions where Nodal signaling is required, including the presumptive mesoderm and endoderm at the blastula and gastrula stages and the prechordal mesoderm at later stages. In Xenopus embryos, overexpression of pitx2 caused ectopic expression of goosecoid and sox-17 and interfered with mesoderm formation. Overexpression of pitx2 in Xenopus animal cap explants partially mimics the effects of Nodal overexpression, suggesting that pitx2 is a mediator of Nodal signaling during specification of the endoderm and prechordal plate, but not during mesoderm induction. We further demonstrate that pitx2 is induced by Nodal signaling in Xenopus animal caps and that the early expression of zebrafish pitx2 is absent when the Nodal signaling pathway is inactive. Inhibition of pitx2 function using a chimeric EnR-pitx2 blocked specification of the mesoderm and endoderm and caused severe embryonic defects resembling those seen when Nodal signaling is inhibited. Following inhibition of pitx2 function, the fate of ventral vegetal blastomeres was shifted from an endodermal to a more mesodermal fate, an effect that was reversed by wild-type pitx2. Finally, we show that inhibition of pitx2 function interferes with the response of cells to Nodal signaling. Our results provide direct evidence that pitx2 function is required for normal specification of the endodermal and mesodermal germ layers.

Amino Acid Sequence↗
PubMed2002

Making worm guts: the gene regulatory network of the Caenorhabditis elegans endoderm.

The nematode Caenorhabditis elegans is a triploblastic ecdysozoan, which, although it contains too few cells during embryogenesis to create discernible germ "layers," deploys similar programs for germ layer differentiation used in animals with many more cells. The endoderm arises from a single progenitor, the E cell, and is selected from among three possible fates by a three-state combinatorial regulatory system involving intersecting cell-intrinsic and intercellular signals. The core gene regulatory cascade that drives endoderm development, extending from early maternal regulators to terminal differentiation genes, is characterized by activation of successive tiers of transcription factors, including a sequential cascade of redundant GATA transcription factors. Each tier is punctuated by a cell division, raising the possibility that intercession of one cell cycle round, or DNA replication, is required for activation of the next tier. The existence of each tier in the regulatory hierarchy is justified by the assignment of a unique task and each invariably performs at least two functions: to activate the regulators in the next tier and to perform one other activity distinct from that of the next tier. While the regulatory inputs that initiate endoderm development are highly divergent, they mobilize a gene regulatory network for endoderm development that appears to be common to all triploblastic metazoans. Genome-wide functional genomic approaches, including identification of >800 transcripts that exhibit the same regulatory patterns as a number of endoderm-specific genes, are contributing to elucidation of the complete endoderm gene regulatory network in C. elegans. Dissection of the architecture of the C. elegans endoderm network may provide insights into the evolutionary plasticity and origins of this germ layer.

Animals↗
PubMed1987

Subcellular organization of the yolk syncytial-endoderm complex in the preimplantation yolk sac of the shark, Rhizoprionodon terraenovae.

The structure of the yolk syncytial-endoderm complex of the preimplantation yolk sac of the shark is examined by light- and transmission electron microscopy. The yolk syncytium is bounded by a membrane that is anchored to the plasmalemma of adjacent endoderm cells by desmosomes. Enlarged nuclei, rough endoplasmic reticulum, Golgi complexes, mitochondria, and other cellular organelles populate the syncytium. Microtubules and filamentous elements are also observed free in the syncytium. Yolk is present as pleomorphic droplets, the profiles of which are generally spherical but may be vesicular, especially at the periphery of large yolk droplets. Occasionally, large yolk droplets have a paracrystalline configuration. Small yolk droplets are modulated through the Golgi complex of the yolk syncytium, and it is suggested that acid hydrolases are added there. Small yolk droplets released from the maturing face of the Golgi complex are sequestered in membrane-limited packets. The membrane of the packets fuses with the membrane enveloping the yolk syncytium and the yolk droplets are released into the yolk syncytial-endoderm interspace. Subsequently, the yolk droplets are endocytosed by the endoderm. Yolk droplets disperse and fuse to form the large irregular yolk inclusions of the endoderm. Yolk metabolites are transported out of the endoderm through the yolk sac endothelium. The yolk sac endoderm thus mediates the transfer of metabolites from the yolk mass to the extraembryonic circulation.

Animals↗
PubMed1989

Is extraembryonic angiogenesis in the chick embryo controlled by the endoderm? A morphology study.

The area vasculosa of the chick embryo is subdivided into two concentric zones: the inner transparent area pellucida vasculosa (AVP) and the less transparent surrounding area opaca vasculosa (AOV). The different optical properties of these zones are caused by the different morphology of the endoderm, which consists of flat cells in the APV and of high-prismatic cells containing large yolk vacuoles in the AOV. The present study describes how this endodermal subdivision of the area vasculosa is related to the development of the extraembryonic vascular pattern. By injection of ink into the vascular system of chick embryos at stages 12 to 20 (Hamburger and Hamilton 1951 "HH"), it has been demonstrated that the vascular net of the area vasculosa from stage 14 (HH) onwards develops into different patterns in APV and AOV. The small loops of uniform capillary vessels of stage 13 (HH) are widened due to the rapid expansion of the extraembryonic mesoderm. In the AOV from stage 14 (HH) onwards numerous small blood vessels sprout into the enlarged intervascular spaces. This process is maximal at stage 17 (HH). In contrast, the blood vessels of the APV remain largely unbranched. These findings suggest that the development of the extraembryonic vascular pattern is controlled by the endodermal pattern. To test this hypothesis, both zones (APV and AOV) were examined by light microscopy, transmission and scanning electron microscopy, in vivo observations and by treatment with bromodeoxyuridine (BrdU). TEM examinations show that the ultrastructural organization of the APV mesoderm is different from that of the AOV: The splanchnopleuric cells of the APV form a continuous cover around the endothelial cells connected by numerous desmosomes, whereas the splanchnopleuric cells of the AOV are frequently separated by gaps. The largest gaps are seen in the small blood vessels at stage 17 (HH). These results should be considered in relation to the dynamic changes in the vascular pattern of the AOV. The endodermal cells of APV and AOV are two different populations. In vivo observation of the endodermal transition from APV to AOV detected no transformations of APV cells into AOV cells or vice versa. The borderline between the zones is stable. The AOV endoderm, having been overgrown by the expanding mesoderm, stops proliferating almost completely, whereas the proliferation of the APV endoderm is unaffected by contact with the mesoderm. The rate of its proliferation is approximately as high as that of the AOV prior to contact with the expanding mesoderm (results after treatment with BrdU).(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1992

Localization of endoderm-specific mRNAs in differentiating F9 embryoid bodies.

Primitive endoderm in the peri-implantation mouse embryo differentiates into two separate lineages, visceral and parietal endoderm (VE and PE). F9 teratocarcinoma cells, when grown in suspension in the presence of retinoic acid (RA), differentiate into embryoid bodies (EBs) which can be used as a model system to study the spatially appropriate induction of VE- and PE-specific gene expression. We have used a whole mount in situ hybridization technique to follow the localization of VE-specific AFP and PE-specific tPA mRNAs during EB differentiation. We show that the putative endoderm-specific markers are localized to the endoderm in mature EBs, and that AFP mRNA is localized to the apical edge of the VE in RA EBs. Surprisingly, prior to localization of endoderm-specific markers at the periphery of EBs, these genes are expressed at low to moderate levels in all cells of the EB. Our data suggest that the establishment of endoderm in EBs is position dependent and not the result of sorting out of predetermined, randomly distributed cells. Our observations also suggest a two-step process for establishing endoderm-specific gene expression, involving up-regulation of transcripts throughout the EB prior to the restriction of their expression to the outer differentiating cell layer.

Animals↗
PubMed2005

Determination of the minimal domains of Mix.3/Mixer required for endoderm development.

The Mix/Bix family of Pax-like homeodomain transcription factors is expressed early in vertebrate development and play important roles in endoderm and mesoderm formation. Like other Pax-related homeodomain proteins, the Mix/Bix family binds DNA as monomers or dimers and dimerization is mediated by the homeodomain. While the Mix/Bix family shares extensive sequence homology within the DNA-binding homeodomain, ectopic expression of these proteins has profoundly different outcomes. Expression of Xenopus Mix.3/Mixer in explanted ectoderm results in endoderm differentiation, whereas Mix.1 expression does not. In this study we sought to define the domains of Mix.3/Mixer that are responsible for this endoderm inducing activity. We generated domain swap mutants between Mix.3/Mixer and Mix.1 and tested their ability to induce endoderm in explanted ectoderm. We demonstrate that the homeodomain and sixty-two amino acids in the carboxyl terminus are required to induce endoderm and that these domains must be on the same polypeptide and can not act in trans as a heterodimer. A Smad2 interaction motif in Mix.3/Mixer is involved in endoderm differentiation but is not essential. Thus, we have defined the regions of Mix.3/Mixer that confer endoderm-inducing activity. These studies reveal a novel co-operation between the homeodomain and a small domain in the carboxyl terminal region that is essential for Mix.3/Mixer function.

Animals↗
PubMed2005

Fate and plasticity of the endoderm in the early chick embryo.

In vertebrates, the endoderm is established during gastrulation and gradually becomes regionalized into domains destined for different organs. Here, we present precise fate maps of the gastrulation stage chick endoderm, using a method designed to label cells specifically in the lower layer. We show that the first population of endodermal cells to enter the lower layer contributes only to the midgut and hindgut; the next cells to ingress contribute to the dorsal foregut and followed finally by the presumptive ventral foregut endoderm. Grafting experiments show that some migrating endodermal cells, including the presumptive ventral foregut, ingress from Hensen's node, not directly into the lower layer but rather after migrating some distance within the middle layer. Cell transplantation reveals that cells in the middle layer are already committed to mesoderm or endoderm, whereas cells in the primitive streak are plastic. Based on these results, we present a revised fate map of the locations and movements of prospective definitive endoderm cells during gastrulation.

Animals↗
PubMed2004

The transcription factor hepatocyte nuclear factor-6/Onecut-1 controls the expression of its paralog Onecut-3 in developing mouse endoderm.

During development, the endoderm gives rise to several organs, including the pancreas and liver. This differentiation process requires spatial and temporal regulation of gene expression in the endoderm by a network of tissue-specific transcription factors whose elucidation is far from complete. These factors include the Onecut protein hepatocyte nuclear factor-6 (HNF-6), which controls pancreas and liver development as shown in our previous work on Hnf6 knock-out embryos. In mammals, HNF-6 has two paralogs, Onecut-2 (OC-2) and OC-3, whose patterns of expression in the adult overlap with that of HNF-6. In the present work, we examine the expression profile of the three Onecut factors in the developing mouse endoderm. We show that HNF-6, OC-2, and OC-3 are expressed sequentially, which defines new steps in endoderm differentiation. By analyzing Hnf6 knock-out embryos we find that HNF-6 is required for expression of the Oc3 gene in the endoderm. We show that OC-3 colocalizes with HNF-6 in the endoderm and in embryonic pancreas and liver. Based on transfection, chromatin immunoprecipitation, and whole embryo electroporation experiments, we demonstrate that HNF-6 can bind to and stimulate the expression of the Oc3 gene. This study identifies a regulatory cascade between two paralogous transcription factors, sheds new light on the interpretation of the Hnf6 knock-out phenotype, and broadens the transcription factors network operating during development of the endoderm, liver, and pancreas.

Animals↗
PubMedDate not supplied

Roles for Rho/ROCK and vinculin in parietal endoderm migration.

The first cell migration event in the mouse embryo is the movement of parietal endoderm cells from the surface of the inner cell mass facing the blastocoel cavity to line the inner surface of the trophectoderm. F9 embryoid bodies provide an in vitro model for this event. They have an inner core of undifferentiated stem cells surrounded by an outer visceral endoderm layer. When plated on a laminin coated substrate, visceral endoderm transitions to parietal endoderm and migrates onto the dish, away from the attached embryoid body. We now show that this outgrowth contains abundant focal complexes and focal adhesions, as well as lamellipodia and filopodia. Treatment with the ROCK inhibitor Y-27632 promotes a 2-fold increase in outgrowth, and a transition from focal adhesions and associated stress fibers, to focal complexes and a decrease in stress fibers. ROCK inhibition also leads to an increase in lamellipodia. Inhibition of RhoA by transfection of a vector encoding C3 transferase, direct administration of the C3 enzyme, or transfection of a vector encoding p190 Rho GTPase Activating Protein also promotes outgrowth and an apparent transition from focal adhesions to focal complexes. Parietal endoderm outgrowth generated using vinculin-deficient F9 stem cells migrates 2-fold further than wild type cultures, but this outgrowth retains the morphology of wild type parietal endoderm, including focal adhesions and stress fibers. Addition of Y-27632 to vinculin-null outgrowth cultures further stimulates migration an additional 2-fold, supporting the conclusion that Rho/ROCK and vinculin regulate parietal endoderm outgrowth by distinct pathways.

ADP Ribose Transferases↗
PubMed1990

Intestinal mesenchyme provokes differentiation of intestinal endocrine cells in gizzard endoderm.

The gizzard (muscular stomach) of chicks is deficient in endocrine cells at hatching. It has previously been shown that proventricular types and proportions of endocrine cells can be induced in gizzard endoderm under the influence of proventricular (glandular stomach) mesenchyme. In order to test its capacity to form nongastric endocrine cell types, gizzard endoderm of 3.75- to 5-day chick embryos was combined with mesenchyme from the small intestine of 3.5- to 4-day quail embryos. The combinations were grown as chorio-allantoic grafts until they attained an incubation age comparable to that of hatching chicks. Controls comprised reassociated endoderm and mesenchyme of chick gizzard and of quail intestine. In the experimental grafts, morphogenesis was predominantly intestinal but some grafts showed gizzard-like features, particularly if the endoderm had been provided by older donors. All intestinal endocrine cell types, including those also found in the normal proventriculus (serotonin-, glucagon-, pancreatic polypeptide-, neurotensin- and somatostatin-immunoreactive cells) differentiated in experimental grafts, some even where morphogenesis was gizzard-like. Hence progenitors of not only gastric, but also intestinal, endocrine cells are indeed present in gizzard endoderm. The possibility that gizzard mesenchyme is inhibitory to endocrine cell differentiation is mooted. Motilin- and secretin-immunoreactive cells, which are characteristic of the intestine but not of the proventriculus of chicks at hatching, were respectively sparse or absent when the endoderm was derived from older donors. Thus the ability of gizzard endoderm to differentiate into nongastric endocrine cell types declines before its capacity to form gastric types. The unexpected appearance of gastrin-releasing peptide (GRP)-immunoreactive cells, a proventricular type not found in normal chick intestine, suggests that the intestinal mesenchyme, at least in this instance, was exercising a permissive role.

Animals↗
PubMed2004

Distinct GATA6- and laminin-dependent mechanisms regulate endodermal and ectodermal embryonic stem cell fates.

This study investigates the establishment of alternative cell fates during embryoid body differentiation when ES cells diverge into two epithelia simulating the pre-gastrulation endoderm and ectoderm. We report that endoderm differentiation and endoderm-specific gene expression, such as expression of laminin 1 subunits, is controlled by GATA6 induced by FGF. Subsequently, differentiation of the non-polar primitive ectoderm into columnar epithelium of the epiblast is induced by laminin 1. Using GATA6 transformed Lamc1-null endoderm-like cells, we demonstrate that laminin 1 exhibited by the basement membrane induces epiblast differentiation and cavitation by cell-to-matrix/matrix-to-cell interactions that are similar to the in vivo crosstalk in the early embryo. Pharmacological and dominant-negative inhibitors reveal that the cell shape change of epiblast differentiation requires ROCK, the Rho kinase. We also show that pluripotent ES cells display laminin receptors; hence, these stem cells may serve as target for columnar ectoderm differentiation. Laminin is not bound by endoderm derivatives; therefore, the sub-endodermal basement membrane is anchored selectively to the ectoderm, conveying polarity to its assembly and to the differentiation induced by it. Unique to these interactions is their flow through two cell layers connected by laminin 1 and their involvement in the differentiation of two epithelia from the same stem cell pool: one into endoderm controlled by FGF and GATA6; and the other into epiblast regulated by laminin 1 and Rho kinase.

Basement Membrane↗
PubMed2005

Endoderm-derived Fgf3 is necessary and sufficient for inducing neurogenesis in the epibranchial placodes in zebrafish.

In vertebrates, epibranchial placodes are transient ectodermal thickenings that contribute sensory neurons to the epibranchial ganglia. These ganglia innervate internal organs and transmit information on heart rate, blood pressure and visceral distension from the periphery to the central nervous system. Despite their importance, the molecular mechanisms that govern the induction and neurogenesis of the epibranchial placodes are only now being elucidated. In this study, we demonstrate that endoderm is required for neurogenesis of the zebrafish epibranchial placodes. Mosaic analyses confirm that endoderm is the source of the neurogenic signal. Using a morpholino knockdown approach, we find that fgf3 is required for the majority of placode cells to undergo neurogenesis. Tissue transplants demonstrate that fgf3 activity is specifically required in the endodermal pouches. Furthermore, ectopic fgf3 expression is sufficient for inducing phox2a-positive neurons in wild-type and endoderm-deficient embryos. Surprisingly, ectodermal foxi1 expression, a marker for the epibranchial placode precursors, is present in both endoderm-deficient embryos and fgf3 morphants, indicating that neither endoderm nor Fgf3 is required for initial placode induction. Based on these findings, we propose a model for epibranchial placode development in which Fgf3 is a major endodermal determinant required for epibranchial placode neurogenesis.

Animals↗
PubMed2006

Retinoids signal directly to zebrafish endoderm to specify insulin-expressing beta-cells.

During vertebrate development, the endodermal germ layer becomes regionalized along its anteroposterior axis to give rise to a variety of organs, including the pancreas. Genetic studies in zebrafish and mice have established that the signaling molecule retinoic acid (RA) plays a crucial role in endoderm patterning and promotes pancreas development. To identify how RA signals to pancreatic progenitors in the endoderm, we have developed a novel cell transplantation technique, using the ability of the SOX32 transcription factor to confer endodermal identity, to selectively target reagents to (or exclude them from) the endodermal germ layer of the zebrafish. We show that RA synthesized in the anterior paraxial mesoderm adjacent to the foregut is necessary for the development of insulin-expressing beta-cells. Conversely, RA receptor function is required in the foregut endoderm for insulin expression, but not in mesoderm or ectoderm. We further show that activation of RA signal transduction in endoderm alone is sufficient to induce insulin expression. Our results reveal that RA is an instructive signal from the mesoderm that directly induces precursors of the endocrine pancreas. These findings suggest that RA will have important applications in the quest to induce islets from stem cells for therapeutic uses.

Animals↗
PubMed2006

Global analysis of the transcriptional network controlling Xenopus endoderm formation.

A conserved molecular pathway has emerged controlling endoderm formation in Xenopus zebrafish and mice. Key genes in this pathway include Nodal ligands and transcription factors of the Mix-like paired homeodomain class, Gata4-6 zinc-finger factors and Sox17 HMG domain proteins. Although a linear epistatic pathway has been proposed, the precise hierarchical relationships between these factors and their downstream targets are largely unresolved. Here, we have used a combination of microarray analysis and loss-of-function experiments to examine the global regulatory network controlling Xenopus endoderm formation. We identified over 300 transcripts enriched in the gastrula endoderm, including most of the known endoderm regulators and over a hundred uncharacterized genes. Surprisingly only 10% of the endoderm transcriptome is regulated as predicted by the current linear model. We find that Nodal genes, Mixer and Sox17 have both shared and distinct sets of downstream targets, and that a number of unexpected autoregulatory loops exist between Sox17 and Gata4-6, between Sox17 and Bix1/Bix2/Bix4, and between Sox17 and Xnr4. Furthermore, we find that Mixer does not function primarily via Sox17 as previously proposed. These data provides new insight into the complexity of endoderm formation and will serve as valuable resource for establishing a complete endoderm gene regulatory network.

Animals↗
PubMed1995

Targeted mutagenesis of the transcription factor GATA-4 gene in mouse embryonic stem cells disrupts visceral endoderm differentiation in vitro.

Transcription factor GATA-4 belongs to a family of zinc finger proteins involved in lineage determination. GATA-4 is first expressed in yolk sac endoderm of the developing mouse and later in cardiac tissue, gut epithelium and gonads. To delineate the role of this transcription factor in differentiation and early development, we studied embryoid bodies derived from mouse embryonic stem (ES) cells in which both copies of the Gata-4 gene were disrupted. Light and electron microscopy demonstrated that embryoid bodies formed from wild-type and heterozygous deficient ES cells were covered with a layer of visceral yolk sac endoderm, whereas no yolk sac endoderm was evident on the surface of the homozygous deficient embryoid bodies. Independently selected homozygous deficient cell lines displayed this distinctive phenotype, suggesting that it was not an artifact of clonal variation. Biochemical markers of visceral endoderm formation, such as alpha-feto-protein, hepatocyte nuclear factor-4 and binding sites for Dolichos biflorus agglutinin, were absent from the homozygous deficient embryoid bodies. Examination of other differentiation markers in the mutant embryoid bodies, studies of ES cell-derived teratocarcinomas and chimeric mouse analysis demonstrated that GATA-4-deficient ES cells have the capacity to differentiate along other lineages. We conclude that, under in vitro conditions, disruption of the Gata-4 gene results in a specific block in visceral endoderm formation. These homozygous deficient cells should yield insights into the regulation of yolk sac endoderm development and the factors expressed by visceral endoderm that influence differentiation of adjoining ectoderm/mesoderm.

Animals↗
PubMed1997

The allocation of early blastomeres to the ectoderm and endoderm is variable in the sea urchin embryo.

During sea urchin development, a tier-to-tier progression of cell signaling events is thought to segregate the early blastomeres to five different cell lineages by the 60-cell stage (E. H. Davidson, 1989, Development 105, 421-445). For example, the sixth equatorial cleavage produces two tiers of sister cells called 'veg1' and 'veg2,' which were projected by early studies to be allocated to the ectoderm and endoderm, respectively. Recent in vitro studies have proposed that the segregation of veg1 and veg2 cells to distinct fates involves signaling between the veg1 and veg2 tiers (O. Khaner and F. Wilt, 1991, Development 112, 881-890). However, fate-mapping studies on 60-cell stage embryos have not been performed with modern lineage tracers, and cell interactions between veg1 and veg2 cells have not been shown in vivo. Therefore, as an initial step towards examining how archenteron precursors are specified, a clonal analysis of veg1 and veg2 cells was performed using the lipophilic dye, DiI(C16), in the sea urchin species, Lytechinus variegatus. Both veg1 and veg2 descendants form archenteron tissues, revealing that the ectoderm and endoderm are not segregated at the sixth cleavage. Also, this division does not demarcate cell type boundaries within the endoderm, because both veg1 and veg2 descendants make an overlapping range of endodermal cell types. The allocation of veg1 cells to ectoderm and endoderm during cleavage is variable, as revealed by both the failure of veg1 descendants labeled at the eighth equatorial division to segregate predictably to either tissue and the large differences in the numbers of veg1 descendants that contribute to the ectoderm. Furthermore, DiI-labeled mesomeres of 32-cell stage embryos also contribute to the endoderm at a low frequency. These results show that the prospective archenteron is produced by a larger population of cleavage-stage blastomeres than believed previously. The segregation of veg1 cells to the ectoderm and endoderm occurs relatively late during development and is unpredictable, indicating that later cell position is more important than the early cleavage pattern in determining ectodermal and archenteron cell fates.

Animals↗
PubMed1998

The transcription factor HNF3beta is required in visceral endoderm for normal primitive streak morphogenesis.

During early embryogenesis, the transcription factor HNF3beta is expressed in visceral and definitive endoderm, node, notochord and floorplate. A targeted mutation in the HNF3&bgr ; gene results in the lack of a definitive node and notochord. Furthermore, lack of HNF3beta results in failure of proper primitive streak elongation. To address whether HNF3beta is required in visceral endoderm, we have used tetraploid embryo-ES cell aggregations to generate chimeric mouse embryos with wild-type visceral endoderm and homozygous mutant HNF3beta embryonic ectoderm or vice versa. Replacing the visceral endoderm of mutant HNF3beta embryos rescued proper primitive streak elongation and, conversely, mutant visceral endoderm imposed a severe embryonic-extraembryonic constriction on wild-type embryonic ectoderm. Restoration of normal streak morphogenesis was not sufficient to allow formation of the node and notochord in HNF3beta mutant embryos. Thus, our results demonstrate that HNF3beta has two separate roles in primitive streak formation. One is to act within the visceral endoderm to promote proper streak morphogenesis. The second is autonomous to the node and its precursors and involves specification of node and notochord cell fates. HNF3beta mutant embryos rescued for the embryonic-extraembryonic constriction developed further than mutant embryos, allowing examination of later roles for HNF3beta. We show that such mutant embryos lack foregut and midgut endoderm. In addition, left-right asymmetry is affected in the mutant embryos.

Animals↗
PubMed1999

Variant hepatocyte nuclear factor 1 is required for visceral endoderm specification.

Genetic and molecular evidence indicates that visceral endoderm, an extraembryonic cell lineage, is required for gastrulation, early anterior neural patterning, cell death and specification of posterior mesodermal cell fates. We show that variant Hepatocyte Nuclear Factor 1 (vHNF1), a homeodomain-containing transcription factor first expressed in the primitive endoderm, is required for the specification of visceral endoderm. vHnf1-deficient mouse embryos develop normally to the blastocyst stage, start implantation, but die soon afterwards, with abnormal or absent extraembryonic region, poorly organised ectoderm and no discernible visceral or parietal endoderm. However, immunostaining analysis of E5.5 nullizygous mutant embryos revealed the presence of parietal endoderm-like cells lying on an abnormal basal membrane. Homozygous mutant blastocyst outgrowths or differentiated embryonic stem cells do not express early or late visceral endoderm markers. In addition, in vHnf1 null embryoid bodies there is no activation of the transcription factors HNF-4alpha1, HNF1alpha and HNF-3gamma. Aggregation of vHnf1-deficient embryonic stem cells with wild-type tetraploid embryos, which contribute exclusively to extraembryonic tissues, rescues periimplantation lethality and allows development to progress to early organogenesis. Our results place vHNF1 in a preeminent position in the regulatory network that specifies the visceral endoderm and highlight the importance of this cell lineage for proper growth and differentiation of primitive ectoderm in pregastrulating embryos.

Animals↗