hecate, a zebrafish maternal effect gene, affects dorsal organizer induction and intracellular calcium transient frequency

Abstract

A zebrafish maternal effect mutation, in the gene hecate, results in embryos that have defects in the formation of dorsoanterior structures and altered calcium release. hecate mutant embryos lack nuclear accumulation of beta-catenin and have reduced expression of genes specific to the dorsal organizer. We found that hecate mutant embryos exhibit a nearly 10-fold increase in the frequency of intracellular Ca2+ transients normally present in the enveloping layer during the blastula stages. Inhibition of Ca2+ release leads to ectopic expression of dorsal genes in mutant embryos suggesting that Ca2+ transients are important in mediating dorsal gene expression. Inhibition of Ca2+ release also results in the expression of dorsal-specific genes in the enveloping layer in a beta-catenin-independent manner, which suggests an additional function for the Ca2+ transients in this cellular layer. The mutant phenotype can be reversed by the expression of factors that activate Wnt/beta-catenin signaling, suggesting that the Wnt/beta-catenin pathway, at least as activated by an exogenous Wnt ligand, is intact in hec mutant embryos. Our results are consistent with a role for the hecate gene in the regulation of Ca2+ release during the cleavage stages, which in turn influences dorsal gene expression in both marginal cells along the dorsoventral axis and in the enveloping layer.

Analysis of axis induction mutant embryos reveals morphogenetic events associated with zebrafish yolk extension formation

“…Previous work has shown that high levels of bmp activity lead to a reduction in the dorsally and animally oriented movements associated with convergence and axis extension, respectively (Myers et al, 2002a). Consistent with these results, hec mutant embryos have a marked expansion of bmp expression during the gastrula stages (Lyman Gingerich et al, 2005). Thus, the vegetally directed movement of the hec mutant cells before somitogenesis leads to an expansion of the ventroposterior region.…”

“…1c, asterisk). These cells have been described previously in axis-defective embryos caused by either vegetal ablation (Mizuno et al, 1999;Ober and Schulte-Merker, 1999) or maternal-effect mutations (Kelly et al, 2000;Nojima et al, 2004;Lyman Gingerich et al, 2005), but their origin has not been explained. hec mutant embryos with less severe phenotypes also show dorsoanterior defects, although of correspondingly lesser degrees (Fig.…”

“…The identified mutation in hecate acts as a strict maternal effect mutation, such that embryos from homozygous mutant mothers exhibit axis induction defects regardless of their zygotic phenotype (Lyman Gingerich et al, 2005). Embryos from homozygous hec mutant females (referred to as hec mutant embryos) display a variable loss of dorsoanterior structures compared with embryos from wild-type females (wild-type embryos; Fig.…”

“…For ␤-catenin labeling, embryos were fixed and immunostained according to Lyman Gingerich et al (2005). Embryos were incubated overnight in a 1:500 dilution of mouse monoclonal antibody clone 15B8 (Sigma), followed by incubation with a 1:400 dilution of Alexa 488 -labeled rabbit anti-mouse secondary antibody (Molecular Probes).…”

“…The recessive maternal-effect mutation in the gene hecate (hec) was identified in a gynogenesisbased genetic screen for maternal-effect mutants (Pelegri et al, 2004). Embryos from homozygous females lack dorsoanterior structures at 24 hpf due to a lack of activation of the Wnt/␤-catenin signaling pathway thought to specify the zebrafish dorsal organizer (Lyman Gingerich et al, 2005).…”