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J Rossant

Publications and source records attributed to J Rossant.

At least 37 records · Page 2Linked to original sources
PubMed2000

The organizer factors Chordin and Noggin are required for mouse forebrain development.

In mice, there is evidence suggesting that the development of head and trunk structures is organized by distinctly separated cell populations. The head organizer is located in the anterior visceral endoderm (AVE) and the trunk organizer in the node and anterior primitive streak. In amphibians, Spemann's organizer, which is homologous to the node, partially overlaps with anterior endoderm cells expressing homologues of the AVE markers cerberus, Hex and Hesx1. For mice, this raises the question of whether the AVE and node are independent of each other, as suggested by their anatomical separation, or functionally interdependent as is the case in amphibians. Chordin and Noggin are secreted bone morphogenetic protein (BMP) antagonists expressed in the mouse node, but not in the AVE. Here we show that mice double-homozygous mutants that are for chordin and noggin display severe defects in the development of the prosencephalon. The results show that BMP antagonists in the node and its derivatives are required for head development.

Animals↗
PubMed2000

The SH2 tyrosine phosphatase shp2 is required for mammalian limb development.

The tyrosine phosphatase Shp2 is recruited into tyrosine-kinase signalling pathways through binding of its two amino-terminal SH2 domains to specific phosphotyrosine motifs, concurrent with its re-localization and stimulation of phosphatase activity. Shp2 can potentiate signalling through the MAP-kinase pathway and is required during early mouse development for gastrulation. Chimaeric analysis can identify, by study of phenotypically normal embryos, tissues that tolerate mutant cells (and therefore do not require the mutated gene) or lack mutant cells (and presumably require the mutated gene during their developmental history). We therefore generated chimaeric mouse embryos to explore the cellular requirements for Shp2. This analysis revealed an obligatory role for Shp2 during outgrowth of the limb. Shp2 is specifically required in mesenchyme cells of the progress zone (PZ), directly beneath the distal ectoderm of the limb bud. Comparison of Ptpn11 (encoding Shp2)-mutant and Fgfr1 (encoding fibroblast growth factor receptor-1)-mutant chimaeric limbs indicated that in both cases mutant cells fail to contribute to the PZ of phenotypically normal chimaeras, leading to the hypothesis that a signal transduction pathway, initiated by Fgfr1 and acting through Shp2, is essential within PZ cells. Rather than integrating proliferative signals, Shp2 probably exerts its effects on limb development by influencing cell shape, movement or adhesion. Furthermore, the branchial arches, which also use Fgfs during bud outgrowth, similarly require Shp2. Thus, Shp2 regulates phosphotyrosine-signalling events during the complex ectodermal-mesenchymal interactions that regulate mammalian budding morphogenesis.

Animals↗
PubMed2000

Rom-1 is required for rod photoreceptor viability and the regulation of disk morphogenesis.

The homologous membrane proteins Rom-1 and peripherin-2 are localized to the disk rims of photoreceptor outer segments (OSs), where they associate as tetramers and larger oligomers. Disk rims are thought to be critical for disk morphogenesis, OS renewal and the maintenance of OS structure, but the molecules which regulate these processes are unknown. Although peripherin-2 is known to be required for OS formation (because Prph2-/- mice do not form OSs; ref. 6), and mutations in RDS (the human homologue of Prph2) cause retinal degeneration, the relationship of Rom-1 to these processes is uncertain. Here we show that Rom1-/- mice form OSs in which peripherin-2 homotetramers are localized to the disk rims, indicating that peripherin-2 alone is sufficient for both disk and OS morphogenesis. The disks produced in Rom1-/- mice were large, rod OSs were highly disorganized (a phenotype which largely normalized with age) and rod photoreceptors died slowly by apoptosis. Furthermore, the maximal photoresponse of Rom1-/- rod photoreceptors was lower than that of controls. We conclude that Rom-1 is required for the regulation of disk morphogenesis and the viability of mammalian rod photoreceptors, and that mutations in human ROM1 may cause recessive photoreceptor degeneration.

Animals↗
PubMed1999

Neural induction and patterning in the mouse in the absence of the node and its derivatives.

The signals which induce vertebrate neural tissue and pattern it along the anterior-posterior (A-P) axis have been proposed to emanate from Spemann's organizer, which in mammals is a structure termed the node. However, mouse embryos mutant for HNF3 beta lack a morphological node and node derivatives yet undergo neural induction. Gene expression domains occur at their normal A-P axial positions along the mutant neural tubes in an apparently normal temporal manner, including the most anterior and posterior markers. This neural patterning occurs in the absence of expression of known organizer genes, including the neural inducers chordin and noggin. Other potential signaling centers in gastrulating mutant embryos appear to express their normal constellation of putative secreted factors, consistent with the possibility that neural-inducing and -patterning signals emanate from elsewhere or at an earlier time. Nevertheless, we find that the node and the anterior primitive streak, from which the node derives, are direct sources of neural-inducing signals, as judged by expression of the early midbrain marker Engrailed, in explant-recombination experiments. Similar experiments showed the neural-inducing activity in HNF3 beta mutants to be diffusely distributed. Our results indicate that the mammalian organizer is capable of neural induction and patterning of the neural plate, but that maintenance of an organizer-like signaling center is not necessary for either process.

Animals↗
PubMed1999

Multiple developmental roles of VEGF suggested by a LacZ-tagged allele.

Vascular endothelial growth factor (VEGF) is an angiogenic factor and a potent stimulator of microvascular permeability. It is a mitogen specific for endothelial cells. The expression of VEGF and its two receptors, Flk-1 and Flt-1, is pivotal for the proper formation of blood vessels in embryogenesis as shown by gene-targeting experiments. Interestingly, the loss of even a single allele of VEGF led to embryonic lethality between day E9.5 and day E10.5 in the mouse. To assess the role of VEGF during embryonic development we decided to tag VEGF expression with LacZ, by inserting an IRES (internal ribosome entry site)-LacZ reporter cassette into the 3' untranslated region of the gene. This alteration enabled us to monitor VEGF expression throughout embryonic development at single-cell resolution. beta-Galactosidase expression from the altered VEGF locus was first observed prior to gastrulation and was detectable at all stages of vascular development in the embryo. Later, the specific cellular distribution and the level of VEGF expression indicated its pleiotropic role in development. High expression levels seemed to be associated with vasculogenesis and permeability, whereas lower levels were associated with angiogenesis and cell migration. In addition, we found VEGF expression in a subtype of endothelial cells present in the endocardium. We believe that the LacZ-tagged allele we have generated offers a precise means of detecting VEGF expression under a variety of physiological and pathological conditions.

Alleles↗
PubMed1999

Interaction between Notch signalling and Lunatic fringe during somite boundary formation in the mouse.

BACKGROUND: The process of somitogenesis can be divided into three major events: the prepatterning of the mesoderm; the formation of boundaries between the prospective somites; and the cellular differentiation of the somites. Expression and functional studies have demonstrated the involvement of the murine Notch pathway in somitogenesis, although its precise role in this process is not yet well understood. We examined the effect of mutations in the Notch pathway elements Delta like 1 (Dll1), Notch1 and RBPJkappa on genes expressed in the presomitic mesoderm (PSM) and have defined the spatial relationships of Notch pathway gene expression in this region. RESULTS: We have shown that expression of Notch pathway genes in the PSM overlaps in the region where the boundary between the posterior and anterior halves of two consecutive somites will form. The Dll1, Notch1 and RBPJkappa mutations disrupt the expression of Lunatic fringe (L-fng), Jagged1, Mesp1, Mesp2 and Hes5 in the PSM. Furthermore, expression of EphA4, mCer 1 and uncx4.1, markers for the anterior-posterior subdivisions of the somites, is down-regulated to different extents in Notch pathway mutants, indicating a global alteration of pattern in the PSM. CONCLUSIONS: We propose a model for the mechanism of somite border formation in which the activity of Notch in the PSM is restricted by L-fng to a boundary-forming territory in the posterior half of the prospective somite. In this region, Notch function activates a set of genes that are involved in boundary formation and anterior-posterior somite identity.

Animals↗
PubMed1999

A mouse cerberus/Dan-related gene family.

The Xenopus cerberus gene is able to induce ectopic heads in Xenopus embryos. At the time of its identification, cerberus shared significant homology with only one other protein, the putative rat tumor suppressor protein Dan. Sequence analysis has revealed that cerberus and Dan are members of a family of predicted secreted proteins, here called the can family. The identification of a can-family member in the nematode Caenorhabditis elegans, CeCan1, suggests that this family is of ancient origin. In the mouse, there are at least five family members: Cer1, Drm, PRDC, Dan, and Dte. These genes are expressed in patterns that suggest that they may play important roles in patterning the developing embryo. Cer1 marks the anterior visceral endoderm at E6.5. Dte is expressed asymmetrically in the developing node. Dan is first seen in the head mesoderm of early head fold stage embryos and Drm is expressed in the lateral paraxial mesoderm at E8.5. The region of homology shared by these genes, here called the can domain, closely resembles the cysteine knot motif found in a number of signaling molecules, such as members of the TGFbeta superfamily. Epitope-tagged versions of Cer1 show that, unlike in TGFbeta superfamily members, the cysteine knot motif is not processed away from a proprotein. Recent experiments in Xenopus have suggested that cerberus may act as an inhibitor of BMP signaling. To examine this further, the ability of Dan, Cer1, and human DRM to attenuate Bmp4 signaling has been assessed in P19 cells using pTlx-Lux, a BMP-responsive reporter. All three genes are able to inhibit Bmp4 signaling. These data suggest that the different family members may act to modulate the action of TGFbeta family members during development.

Amino Acid Sequence↗
PubMed1999

Distinct neural stem cells proliferate in response to EGF and FGF in the developing mouse telencephalon.

Multipotent, self-renewing neural stem cells reside in the embryonic mouse telencephalic germinal zone. Using an in vitro neurosphere assay for neural stem cell proliferation, we demonstrate that FGF-responsive neural stem cells are present as early as E8.5 in the anterior neural plate, but EGF-responsive neural stem cells emerge later in development in a temporally and spatially specific manner. By separately blocking EGF and FGF2 signaling, we also show that EGF alone and FGF2 alone can independently elicit neural stem cell proliferation and at relatively high cell densities separate cell nonautonomous effects can substantially enhance the mitogen-induced proliferation. At lower cell densities, neural stem cell proliferation is additive in the presence of EGF and FGF2 combined, revealing two different stem cell populations. However, both FGF-responsive and EGF-responsive neural stem cells retain their self-renewal and multilineage potential, regardless of growth factor conditions. These results support a model in which separate, lineage-related EGF- and FGF-responsive neural stem cells are present in the embryonic telencephalic germinal zone.

Animals↗
PubMed1999

A repertoire of differentially expressed transcription factors that offers insight into mechanisms of human cytotrophoblast differentiation.

During human placental development, specialized cells allocated to the extraembryonic lineage (cytotrophoblasts) invade the uterus, anchoring the conceptus to the decidua and tapping a supply of maternal blood. This unusual behavior requires cytotrophoblasts to assume highly specialized characteristics; some are commonly associated with tumor cells, while others are typical of endothelia. Here we investigated the transcriptional mechanisms that control cytotrophoblast differentiation/invasion. Specifically, we examined the cells' expression of a number of transcription factors, at the RNA level, as they differentiated along the invasive pathway in vitro. Since basic helix-loop-helix (bHLH) proteins play important roles in murine trophoblast differentiation, we first examined their expression by cytotrophoblasts. As in murine placental development, expression of the human homologue of Mash-2 was confined to progenitor cells. But expression of Hand-1, which promotes differentiation of murine trophoblast giant cells, was not detected. We also found that cytotrophoblasts upregulated the expression of bHLH/PAS factors that function in adaptive responses to hypoxia, including hEPAS-1, which is expressed primarily in endothelial cells. Quite unexpectedly, we discovered that cytotrophoblasts express high levels of mRNA encoding the human homologue of the Drosophila neuronal fate gene, glial cells missing-1 (gcm-1). We also found evidence of crosstalk between the bHLH and GCM-1 regulatory networks. Together, these results offer insights into the transcriptional mechanisms that govern cytotrophoblast differentiation/invasion. Interestingly, these mechanisms suggest analogies with those that govern differentiation of murine stem cells allocated to both the intra- and extraembryonic lineages.

Animals↗
PubMed1999

Expression of the T-box gene Eomesodermin during early mouse development.

We report the expression pattern of a murine homolog of the Xenopus laevis T-box gene Eomesodermin. mEomes expression is first detected in the extra-embryonic ectoderm prior to gastrulation, and persists there until head-fold stages. In the embryo proper, mEomes is expressed throughout the early primitive streak, nascent mesoderm and in the anterior visceral endoderm. Although mEomes expression disappears from the embryo at late-streak stages, a second domain of mEomes expression is observed in the telencephalon beginning around E10.5.

Animals↗
PubMed1999

Parental origin-specific expression of Mash2 is established at the time of implantation with its imprinting mechanism highly resistant to genome-wide demethylation.

The Mash2 gene encodes a basic helix-loop-helix transcription factor, which is highly expressed in diploid trophoblast cells of the postimplantation mouse embryo and is required for development of the spongiotrophoblast in order to form a functional placenta. Genomic imprinting of Mash2 has been previously reported; transcriptional inactivation of the paternal wild-type allele in heterozygotes carrying a maternal null allele results in a null-equivalent embryonic lethal phenotype. In order to study the Mash2 imprinting mechanism, we have created a new allele at this locus carrying a targeted insertion of an IRES (internal ribosome entry site)-lacZ cassette within the 3' untranslated region of the gene (referred to as "Mash2-lacZ"). This new allele has made it feasible to monitor paternal Mash2 expression in a wild-type-equivalent background. Our data suggest that parental origin-specific expression of Mash2 begins in the early postimplantation conceptus (5.5 dpc) at the time when trophoblast-specific expression is observed. We also show that the paternal allele is continuously repressed up to 9.5 dpc in the developing ectoplacental cone (EPC) and early chorio-allantoic placenta, with some cells escaping paternal repression. When maternally inherited, lacZ expression from this allele reflects the expression pattern of endogenous Mash2 transcripts up to 8.5 dpc. Furthermore, we have addressed the question of a requirement for DNA methylation for the Mash2 imprinting mechanism by crossing our Mash2-lacZ mice with mice mutant for Dnmt1 (DNA-methyltransferase1). Our results show a partial loss of transcriptional repression of the paternal allele in Dnmt1 deficient background. Interestingly, however, this is not sufficient to eliminate the highly biased parental allele-specific expression of Mash2. Thus, the preferential maternal expression of the gene is still maintained in Dnmt1 null mutant embryos, although methylation analyses demonstrate that the Mash2 locus is highly demethylated in Dnmt1 null mutant embryos. The locus is also highly demythyled in wild-type EPCs. Our results suggest the possibility that a mechanism other than DNA methylation, such as allele-specific chromatin conformation, may be involved in maintenance of parental origin-specific expression of Mash2.

Animals↗
PubMed1999

Interaction of the TEK and TIE receptor tyrosine kinases during cardiovascular development.

TEK (TIE2) and TIE (TIE1) are structurally related receptor tyrosine kinases expressed in endothelial cells and their precursors. Genetic studies in the mouse have revealed essential functions of both receptors in angiogenic expansion of the vasculature during development. As previously shown, mouse embryos homozygous for a disrupted Tek allele die by day 10.5 of embryogenesis due to endocardial defects, hemorrhaging, and impaired vascular network formation. Furthermore, TIE is required cell autonomously for endothelial cell survival and extension of the vascular network during late embryogenesis. Here we have investigated possible redundancy in the TEK and TIE signalling pathways during vascular development. Vasculogenesis proceeds normally in embryos lacking both TEK and TIE, although such embryos die early in gestation of multiple cardiovascular defects. Mosaic analysis revealed an absolute requirement for TEK in the endocardium at E10.5, whereas TEK and TIE are dispensable for the initial assembly of the rest of the vasculature. In contrast, both receptors are required in the microvasculature during late organogenesis and in essentially all blood vessels of the adult. This analysis demonstrates essential functions for TEK and TIE in maintaining the integrity of the mature vasculature.

Animals↗
PubMed1999

The basic-helix-loop-helix protein pod1 is critically important for kidney and lung organogenesis.

Epithelial-mesenchymal interactions are required for the development of all solid organs but few molecular mechanisms that underlie these interactions have been identified. Pod1 is a basic-helix-loop-helix (bHLH) transcription factor that is highly expressed in the mesenchyme of developing organs that include the lung, kidney, gut and heart and in glomerular visceral epithelial cells (podocytes). To determine the function of Pod1 in vivo, we have generated a lacZ-expressing null Pod1 allele. Null mutant mice are born but die in the perinatal period with severely hypoplastic lungs and kidneys that lack alveoli and mature glomeruli. Although Pod1 is exclusively expressed in the mesenchyme and podocytes, major defects are observed in the adjacent epithelia and include abnormalities in epithelial differentiation and branching morphogenesis. Pod1 therefore appears to be essential for regulating properties of the mesenchyme that are critically important for lung and kidney morphogenesis. Defects specific to later specialized cell types where Pod1 is expressed, such as the podocytes, were also observed, suggesting that this transcription factor may play multiple roles in kidney morphogenesis.

Alleles↗
PubMed1998

Promotion of trophoblast stem cell proliferation by FGF4.

The trophoblast cell lineage is essential for the survival of the mammalian embryo in utero. This lineage is specified before implantation into the uterus and is restricted to form the fetal portion of the placenta. A culture of mouse blastocysts or early postimplantation trophoblasts in the presence of fibroblast growth factor 4 (FGF4) permitted the isolation of permanent trophoblast stem cell lines. These cell lines differentiated to other trophoblast subtypes in vitro in the absence of FGF4 and exclusively contributed to the trophoblast lineage in vivo in chimeras.

Animals↗
PubMed1998

Opposite phenotypes of hypomorphic and Y766 phosphorylation site mutations reveal a function for Fgfr1 in anteroposterior patterning of mouse embryos.

Intercellular communication is needed for both the generation of the mesodermal germ layer and its division into distinct subpopulations. To dissect the functions of fibroblast growth factor receptor-1 (FGFR1) during mouse gastrulation as well as to gain insights into its possible roles during later embryonic development, we have introduced specific mutations into the Fgfr1 locus by gene targeting. Our results show functional dominance of one of the receptor isoforms and suggest a function for the autophosphorylation of site Y766 in the negative regulation of FGFR1 activity. Y766F and hypomorphic mutations in Fgfr1 generate opposite phenotypes in terms of homeotic vertebral transformations, suggesting a role for FGFR1 in patterning the embryonic anteriorposterior axis by way of regulation of Hox gene activity.

Alleles↗
PubMed1998

Regulation of a transcription factor network required for differentiation and metabolism.

Hepatocyte nuclear factors (HNFs) are a heterogeneous class of evolutionarily conserved transcription factors that are required for cellular differentiation and metabolism. Mutations in HNF-1alphaand HNF-4alpha genes impair insulin secretion and cause type 2 diabetes. Regulation of HNF-4/HNF-1 expression by HNF-3alpha and HNF-3beta was studied in embryoid bodies in which one or both HNF-3alpha or HNF-3beta alleles were inactivated. HNF-3beta positively regulated the expression of HNF-4alpha/HNF-1alpha and their downstream targets, implicating a role in diabetes. HNF-3beta was also necessary for expression of HNF-3alpha. In contrast, HNF-3alpha acts as a negative regulator of HNF-4alpha/HNF-1alpha demonstrating that HNF-3alpha and HNF-3beta have antagonistic transcriptional regulatory functions in vivo. HNF-3alpha does not appear to act as a classic biochemical repressor but rather exerts its negative effect by competing for HNF-3 binding sites with the more efficient activator HNF-3beta. In addition, the HNF-3alpha/HNF-3beta ratio is modulated by the presence of insulin, providing evidence that the HNF network may have important roles in mediating the action of insulin.

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