Pou5f1 contributes to dorsoventral patterning by positive regulation of vox and modulation of fgf8a expression

Abstract

Pou5f1/Oct-4 in mice is required for maintenance of embryonic pluripotent cell populations. Zebrafish pou5f1 maternal-zygotic mutant embryos (spiel ohne grenzen; MZspg) lack endoderm and have gastrulation and dorsoventral patterning defects. A contribution of Pou5f1 to the control of bmp2b, bmp4 and vox expression has been suggested, however the mechanisms remained unclear and are investigated in detail here. Low-level overexpression of a Pou5f1-VP16 activator fusion protein can rescue dorsalization in MZspg mutants, indicating that Pou5f1 acts as a transcriptional activator during dorsoventral patterning. Overexpression of larger quantities of Pou5f1-VP16 can ventralize wild-type embryos, while overexpression of a Pou5f1-En repressor fusion protein can dorsalize embryos. Lack of Pou5f1 causes a transient upregulation of fgf8a expression after mid-blastula transition, providing a mechanism for delayed activation of bmp2b in MZspg embryos. Overexpression of the Pou5f1-En repressor induces fgf8, suggesting an indirect mechanism of Pou5f1 control of fgf8a expression. Transcription of vox is strongly activated by Pou5f1-VP16 even when translation of zygotically expressed transcripts is experimentally inhibited by cycloheximide. In contrast, bmp2b and bmp4 are not activated under these conditions. We show that Pou5f1 binds to phylogenetically conserved Oct/Pou5f1 sites in the vox promoter, both in vivo (ChIP) and in vitro. Our data reveals a set of direct and indirect interactions of Pou5f1 with the BMP dorsoventral patterning network that serve to fine-tune dorsoventral patterning mechanisms and coordinate patterning with developmental timing.

Pou2, a class V POU-type transcription factor in zebrafish, regulates dorsoventral patterning and convergent extension movement at different blastula stages

“…The involvement of pou2 in DV patterning was previously reported based on the analyses of M-and/or Z-spg mutants (Reim and Brand, 2006) and the effects of mRNA injection (Belting et al, 2011). In the present study we further showed that pou2 is also involved in CE movement.…”

“…An active and repressive form of pou2 was recently reported to cause ventralization and dorsalization, respectively, by mRNA injection (Belting et al, 2011), although the activity of wild-type pou2 was not addressed directly. In the present study, we first sought to reexamine the effects of wild-type pou2 as compared with those of active and repressive pou2 by mRNA injection.…”

Split top: A maternal cathepsin B that regulates dorsoventral patterning and morphogenesis

“…Expansion of gsc at sphere stage shows that dorsoventral patterning is altered prior to the first observable morphological defects in split top mutants. Although not observed in BMP pathway mutants, a similar expansion of dorsal midline mesoderm is observed in other maternal-effect dorsalized mutants, including ints6 (Kapp et al, 2013) and maternal-zygotic (MZ) pou5f3 (formerly pou5f1 or oct4) (Belting et al, 2011;Reim and Brand, 2006).…”

The Vox mRNA and protein expression in zebrafish Pou5f3 MZspg mutant embryos

“…Pou5f3 in the embryo acts as an activator of gene expression determining ventral fates by direct activation of the Vox promoter, and this is a phylogenetically conserved mechanism. In wild type embryos, the Vox expression was enhanced by the Pou5f3, but it was partially suppressed in the mutants (10). However,…”