Fig. 5

wnt signaling regulates endoderm development prior to liver specification. Heat-shock inducible wnt8-GFP or control embryos were incubated for 20 mins at 38 °C at various time points between 50% epiboly and 48 hpf, and were analyzed for alterations in liver size at 72 hpf by in situ hybridization for lfabp. (A, B) Induction of wnt8 at 1 somite (10 hpf) led to grossly abnormal embryos, with cardiac edema and absent livers (49 altered/ -52 scored). (C) Induction of wnt8 at 10 somites (14 hpf) resulted in a markedly enlarged livers compared to controls (141/173). (D–F) foxa3 expression at 24 hpf following heat-shock at 1 somite (10 hpf) revealed a failure of cells to converge at the midline and expand in the a–p direction (36/45); heat-shock at 10 somites (14 hpf) led to enhanced foxa3 expression (52/67). (G–I) Expression of foxa3 at 48 hpf revealed a failure to form organized liver and pancreatic buds after wnt8 induction at 1 somite (10 hpf) (45/61); wnt8 induction at 10 somites (14 hpf) led to an increased liver and decreased pancreatic anlage (52/78). (J) qPCR analysis of the expression levels of wnt target (cyclind1, cmyc), endodermal (foxa3, sox17), hepatic (hhex, prox1) and pancreatic (pdx1) progenitor genes at 48 hpf in controls (red) and following wnt8 induction at either 1 somite (10 hpf; green) or 10 somites (14 hpf; blue). All changes were statistically significant compared to controls (ANOVA, n = 10, p < 0.05).