Placental vascularisation requires the AP-1 component fra1.

Fra1 is an immediate-early gene encoding a member of the AP-1 transcription factor family, which has diverse roles in development and oncogenesis. To determine the function of Fra1 in mouse development, the gene was inactivated by gene targeting. Foetuses lacking Fra1 were severely growth retarded and died between E10.0 and E10.5, owing to defects in extra-embryonic tissues. The placental labyrinth layer, where X-gal staining revealed expression of Fra1, was reduced in size and largely avascular, owing to a marked decrease in the number of vascular endothelial cells, as shown by the lack of Flk1 expression. In contrast, the spongiotrophoblast layer was unaffected and expressed the marker genes 4311 (Tpbp) and Flt1. Furthermore, mutant foetuses exhibited yolk-sac defects that may contribute to their growth retardation and lethality. Importantly, when the placental defect was rescued by injection of Fra1(-)(/)(-) ES cells into tetraploid wild-type blastocysts, Fra1(-)(/)(-) pups were obtained that were no longer growth retarded and survived up to 2 days after birth without apparent phenotypic defects. These data indicate that a defect in the extra-embryonic compartment is causal to the observed lethality, and suggest that Fra1 plays a crucial role in establishing normal vascularisation of the placenta.

Expression of the RNA recognition motif-containing protein SEB-4 during Xenopus embryonic development.

RNA binding proteins play key roles in the post-transcriptional regulation of gene expression. Here we present the molecular cloning and spatio-temporal expression of Xseb-4, which codes for a putative RNA binding protein containing a single RNA recognition motif (RRM). XSEB-4 shares 60-65% identity with the mammalian SEB-4 proteins. Xseb-4 is strongly expressed maternally. Zygotic transcription is initiated in the early gastrula embryo in paraxial mesoderm that is fated to give rise to somites. During the course of gastrulation and neurulation Xseb-4 expression in somitic paraxial mesoderm is centered within the XmyoD expression domain. As development proceeds Xseb-4 expression is in addition initiated in the cardiac primordium and the lens vesicle. In the heart expression is confined to the myocardium. Thus, the RRM-containing putative RNA binding protein XSEB-4 is differentially expressed during embryonic development in Xenopus.

Neuroectodermal specification and regionalization of the Spemann organizer in Xenopus.

During gastrulation in Xenopus convergence and extension movements, mediated by mediolateral intercalations, are the driving force for early neural plate morphogenesis. Here we show that the winged helix transcriptional regulator, Xfd-12' is dynamically expressed in medial neural plate precursors that undergo convergence and extension movements. These medial neuraxial progenitors are specified in and beyond the Spemann organizer prior to specification of the basal anlage of the neural plate. The initiation of Xfd-12' expression coincides with the induction of mesendoderm by Nodal-related growth factors at the late blastula stage. Comparative expression analysis suggests that cellular rearrangements at the pre-gastrulation stage account for regionalization of the Spemann organizer into head and trunk organizer compartments, the latter in which medial neural plate progenitors reside. While the maintenance of Xfd-12' expression in the dorsal non-involuting marginal zone requires FGF signalling, its subsequent positioning along the medial aspect of the neuraxis depends on signalling by Wnt and Nodal-related family members. Based on these findings we propose that XFD-12' is a trunk organizer component that might control convergence and extension movements of medial neural plate precursors during gastrulation.

Cooperation of Pax2 and Pax5 in midbrain and cerebellum development.

Midbrain and cerebellum development depends on an organizing center that is located at the midbrain-hindbrain junction of the vertebrate embryo. Expression of the two closely related transcription factors Pax2 and Pax5 overlaps spatially and temporally in this region of the developing central nervous system. To study a possible interaction of these transcription factors in midbrain and cerebellum patterning, we have generated Pax5, Krd double mutant mice. The transgene-induced Krd mutation corresponds to an approximately 7-centimorgan chromosome 19 deletion that eliminates the entire Pax2 locus. The heterozygous Krd mutation deleting one Pax2 allele had no effect on midbrain and cerebellum development. Moreover, only minor developmental defects were previously observed at the midline of the inferior colliculus and anterior cerebellum in mice that were homozygous for a targeted Pax5 mutation. Similar morphological alterations were observed in 80% of all compound heterozygous Pax5 (+/-) Krd (+/-) mice. However, in the remaining 20% of compound heterozygotes, the inferior colliculi were missing, and the vermis of the cerebellum was severely disrupted due to the failure of the cerebellar primordia to fuse at the midline. Inactivation of the second Pax5 allele in Pax5 (-/-) Krd (+/-) mice resulted in complete loss of the posterior midbrain and cerebellum, as the tissue originating from the midbrain-hindbrain boundary region was deleted in the embryo as early as day 9.5. On the basis of these data, we propose that the cooperation of Pax2 and Pax5 is essential for normal functioning of the organizing center at the midbrain-hindbrain junction.

Polyoma middle T-induced vascular tumor formation: the role of the plasminogen activator/plasmin system.

The middle T antigen of murine Polyomavirus (PymT) rapidly transforms endothelial cells, leading to the formation of vascular tumors in newborn mice. Transformed endothelial (End.) cell lines established from such tumors exhibit altered proteolytic activity as a result of increased expression of urokinase-type plasminogen activator (uPA) and are capable of inducing vascular tumors efficiently when injected into adult mice. In this study we have used mice lacking components of the PA/plasmin system to analyze the role of this system in the transformation process and in tumor growth. We found that the proteolytic status of the host is not a critical determinant for PymT-induced vascular tumor formation. In addition, the lack of either uPA or tissue-type PA (tPA) activity is not limiting for the establishment and proliferation of End. cells in vitro, although the combined loss of both PA activities leads to a marked reduction in proliferation rates. Furthermore, the in vitro morphogenetic properties of mutant End. cells in fibrin gels could only be correlated with an altered proteolytic status in cells lacking both uPA and tPA. However, in contrast with tumors induced by PymT itself, the tumorigenic potential of mutant and wild-type End. cell lines was found to be highly dependent on the proteolytic status of both the tumor cells and the host. Thus, genetic alterations in the PA/plasmin system affect vascular tumor development, indicating that this system is a causal component in PymTmediated oncogenesis.

Complete block of early B cell differentiation and altered patterning of the posterior midbrain in mice lacking Pax5/BSAP.

The Pax5 gene, coding for the transcription factor BSAP, was mutated in the mouse germline by targeted disruption. Homozygous mutant mice were born alive, became growth retarded, and usually died within three weeks. About 5% of mutants survived to adulthood and were fertile, but severely runted. Morphogenesis of the posterior midbrain was affected as early as embryonic day 16.5, leading to a reduction of the inferior colliculus near the midline and to altered foliation of the anterior cerebellum. Moreover, all mutants failed to produce small pre-B, B, and plasma cells owing to a complete arrest of B cell development at an early precursor stage. These data define a key role for Pax5 in early B lymphopoiesis and midbrain patterning.