Wnt11 family dependent morphogenesis during frog gastrulation is marked by the cleavage furrow protein anillin

Abstract

Wnt11 family proteins are ligands that activate a type of Dishevelled-mediated, non-canonical Wnt signaling pathway. Loss of function causes defects in gastrulation and/or anterior-posterior axis extension in all vertebrates. Non-mammalian vertebrate genomes encode two Wnt11 family proteins whose distinct functions have been unclear. We knocked down zygotic Wnt11b and Wnt11, separately and together, in Xenopus laevis. Single morphants exhibited very similar phenotypes of delayed blastopore closure, but they had different phenotypes at the tailbud stage. In response to their very similar gastrulation phenotypes, we chose to characterize dual morphants. Using dark field illuminated time-lapse imaging and kymograph analysis, we identified a failure of dorsal blastopore lip maturation that correlated with slower blastopore closure and failure to internalize the endoderm at the dorsal blastopore lip. We connected these externally visible phenotypes to cellular events in the internal tissues – including the archenteron – by imaging intact embryos stained for anillin and microtubules. The cleavage furrow protein anillin provided an exceptional cytological marker for blastopore lip and archenteron morphogenesis and the consequent disruption through loss of Wnt11 signaling. These cytological changes suggest a novel role for the regulation of contractility and stiffness of the epithelial cells that result in dramatic shape changes and are important in gastrulation.

Maternal Wnt11b regulates cortical rotation during Xenopus axis formation: analysis of maternal-effect wnt11b mutants

“…1 E, time 07 h:00 min:00 s). Similar results were seen using antisense morpholino oligos against all wnt11 family transcripts ( Van Itallie et al, 2022 preprint), suggesting that normal expression of either maternal (of which Wnt11b is the sole contributor) or pan-zygotic Wnt11 proteins is required for timely and complete gastrulation. Because gastrulation involves coordinated regulation of the cytoskeleton, we examined cytoskeletal organisation in +/Z and M/Z embryos.…”

“…One implication of these findings is that Wnt11b would be signalling in a para/autocrine fashion, acting on the single-cell egg to modulate microtubule dynamics (directly or indirectly). Recent proteomic data support the idea that Wnt11b protein expression is elevated during oocyte maturation ( Van Itallie et al, 2022 preprint; Peshkin et al, 2019 preprint – visualised on Xenbase, ) and other studies indicate active translation in the egg (M. D. Sheets, personal communication).…”

Maternal Wnt11b regulates cortical rotation duringXenopusaxis formation: analysis of maternal-effectwnt11bmutants

“…1E, time 07hr:00min:00sec). Similar results were seen using antisense Morpholino oligos against all wnt11 family transcripts (Van Itallie et al, 2022), suggesting that normal expression of either maternal (of which Wnt11b is the sole contributor) or pan zygotic Wnt11 proteins is required for timely and complete gastrulation.…”

“…Recent proteomic data support the idea that Wnt11b protein expression is elevated during oocyte maturation (Van Itallie et al, 2022;Peshkin et al, 2019 -visualised on Xenbase) and other studies indicate active translation in the egg (Michael D. Sheets, personal communication). This pattern is mirrored by Fzd7, a Wnt11 family receptor, suggesting that this signalling pathway becomes active in the egg.…”

“…We note that gastrulation in MZ wnt11b-/- embryos does seem to progress once zygotic expression of wnt11 (wnt11r ) reaches normal levels (∼ stage 13) even though wnt5 homologues are expressed before this time. However, double Morpholino-based knockdown of Wnt11b/Wnt11 results in similarly delayed gastrulation, with abnormal extension of the archenteron (Van Itallie, et al, 2022). Thus, low levels of Wnt11 family function (whether through delayed wnt11 expression in M/Z mutants or through residual protein in knockdowns), with or without Wnt5 function, might be sufficient for ultimate closure of the blastopore.…”