P118. Multiple and stage-specific functions of a Class V POU transcription factor Pou2/Pou5f1 in zebrafish embryos revealed by the transgenic approach

Zebrafish pou2 is considered to be involved in dorsoventral patterning and endoderm formation in early embryos, and in regionalization of the prospective midbrain and hindbrain in the neural plate. Structural analysis and synteny relationship suggested that pou2 is closely related to mouse Oct3/4 , which is the main regulatory gene maintaining the undifferentiated and pluripotent state in embryonic cells and in ES cells. In the present study, we examined the effects of over-expressing wild type or genetically modified pou2 , and obtained the following results. (1) Overexpression of pou2 by mRNA injection affected the convergence and extension (C&E) movement in the neuroectoderm and axial mesoderm during epiboly. At later stages, pou2 gave rise to undulated hindbrain with reduced ventricles, but brain regionalization took place normally. (2) Overexpression of an activated form of Pou2 (VP-pou2) disrupted the entire brain, especially forebrain and midbrain, as well as abolished the axial mesoderm. When a repressive form of Pou2 (EnR-pou2) was overexpressed by mRNA injection, embryos were dorsalized during epiboly, and their isthmus and hindbrain were deformed at later stages. (3) Overexpression of heat-inducible EnR-pou2 gene in embryos elicited different phenotypes depending on the stage of heat induction. EnR-pou2 overexpression dorsalized embryos when induced at the mid-blastula stage, affected C&E movement during epiboly when induced at the late blastula stage, and disrupted the isthmic region when induced around the end of epiboly/gastrulation. WMISH suggested that pax2a is the direct target gene of pou2 at the isthmus. These results together show pleiotropic and stage-specific functions of pou2 in zebrafish embryos from the blastula through the somitogenesis stage in embryonic patterning as well as in neural development.