|Ph.D Student||Snir Mirit|
|Subject||The Xenopus POU91 Protein Regulates Cellular Competence|
during Early Development
|Department||Department of Medicine||Supervisor||Professor Dale Frank|
This research explores the role of XLPOU91, a POU-homeobox protein that regulates the competence transition from mesoderm to neural cell fate. Cellular competence is defined as a cell’s ability to respond to signaling cues as a function of time. In Xenopus laevis, cellular responsiveness to FGF changes during development. At blastula stages, FGF induces mesoderm, but at gastrula stages FGF regulates neuroectoderm formation. A Xenopus Oct3/4 homologue gene, XLPOU91 regulates mesoderm to neuroectoderm competence transitions. Ectopic XLPOU91 expression in Xenopus embryos inhibits FGF induction of Brachyury (XBra), eliminating posterior mesoderm, while neural induction is unaffected. We used a XLPOU91 antisense morpholino oligonucleodide to knock down endogenous XLPOU91 protein activity in Xenopus laevis embryos. Morphant embryos gastrulated slower with blastopore closure occurring at neurula stages. Morphant embryos lagged behind being at early gastrula stage while control embryos were at early neurula stages; morphant embryos had high proliferation in comparison to control embryos. In morphant ectoderm explants, mesoderm responsiveness to FGF was extended from blastula to gastrula stages. In morphant gastrula stage embryos, initial endoderm and mesoderm induction appeared normal. In gastrula stage embryos, the knockdown of XLPOU91 activity induced higher levels of mesodermal XBra expression. In contrast to initial mesoderm and endoderm induction, neural induction was abolished. At later stages, expression of differentiation markers of all three germ layers was highly impaired. Churchill (Chch) and SIP1, two genes regulating neural competence, were not expressed in morphant embryos, suggesting that XLPOU91 lies upstream of these genes. Ectopic XLPOU91 protein level activates Churchill and SIP gene expression in embryos. Ectopic SIP1 or Churchill expression rescued the XLPOU91 morphant phenotype. Therefore, XLPOU91 regulates the FGF competence transition, which controls Chch/SIP1 gene expression that is critical for neural induction. Thus, in the absence of XLPOU91 activity, the time cues driving proper embryonic cell fate formation are lost. In the absence of the XLPOU91/Churchill/SIP1 network, neural induction is impaired and subsequent normal embryonic development is impaired.