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Figure 4


Fig. 4. Wnt11 is required for induction of cardiomyogenesis by GATA4 and GATA6. RT-PCR analysis of cardiomyogenic gene expression (MLC2, TnIc) at control stage 32 (A,C,E) and analysis of cardiomyocyte differentiation (detectable rhythmic beating) at control stage 45 (B,D,F). (A) Xenopus animal cap explants that were injected with GATA4GR or GATA6GR mRNA (G4GR, G6GR, 100 pg), alone or in combination with Wnt11 morpholino (11MO) as indicated, and cultured in the absence or in the presence of dexamethasone from stage 8. Induction of cardiomyogenic gene expression by G4GR and G6GR is abolished by inhibition of Wnt11. (B) Numerical analysis of spontaneous rhythmic beating in animal cap explants injected with G4GR or G6GR mRNA (500 pg or 1000 pg and activated with dexamethasone at stage 8, as indicated). Only G4GR mRNA is capable of inducing rhythmic beating in animal cap explants. (C) Analysis of cardiomyogenic gene expression in sibling animal cap explants of those used for cardiomyocyte differentiation in D, injected with G4GR (500 pg, activated at stage 8) with 11MO as indicated. (D) Numerical analysis of spontaneous rhythmic beating in sibling animal cap explants. Under these conditions (G4GR mRNA, 500 pg), inhibition of Wnt11 reduces, but does not abolish, cardiomyogenic gene expression, consistent with reduced cardiomyocyte differentiation in explants. (E) Cardiomyogenic gene expression in sibling dorsal marginal zone explants (DMZ) or in sibling whole embryos (W/E) of those used for phenotypic analysis in F, which were injected with 11MO as indicated. (F) Numerical analysis of rhythmic beating in DMZ explants and whole embryos, as in E. Morpholino-mediated inhibition of Wnt11 causes strong but not complete reduction of cardiomyogenic gene expression, consistent with the decreased cardiomyogenesis in both DMZ explants and whole embryos. In B,D, the results of three different experiments are combined but in F only the result of one experiment is presented with n representing the number of explants or embryos scored. VMZ, ventral marginal zone.