Fig. 5.

Fig. 5.

The activity of Wnt9a/b does not depend on the endocardium and prevents premature cardiac differentiation in the heart cone. (A-F) Maximum projections of confocal z-scans with ventral views of cardiomyocyte progenitor cells. In the wildtype (A), cardiomyocyte progenitor cells complete fusion into the heart cone by 20 hpf and undergo leftward jogging by 24 hpf (B). In klf2a/b double morphants (C), cardiomyocyte progenitor cells are not defective during the fusion of the cardiac cone (n=10/10 morphants analyzed) or during leftward jogging (D; n=9/9 morphants analyzed). In npas4l morphants (E), which lack all endocardial cells, cardiomyocyte progenitor cells do not fuse correctly at the embryonic midline (n=8/8 embryos analyzed). Yet, the heart cone undergoes leftward jogging (F; n=10/10 embryos analyzed). (G-J) Maximum projections of confocal z-scans of 48 hpf zebrafish hearts. Different to the looped wild-type heart (G), the npas4l morphant heart (H) is strongly ballooned. In comparison, the wnt9a/b double morphant mutant heart (I) is collapsed. The knockdown of wnt9a/b in npas4l^m378 mutants produces the wnt9a/b double morphant cardiac phenotype with a collapsed heart (J; n=30/30 embryos analyzed). This suggests that the wnt9a/b morphant cardiac phenotype is not due to endocardial defects. (K) Quantifications of changes of mRNA expression levels of myocardial differentiation markers by quantitative real-time PCR. Shown is the comparison in Tg(hsp70l:wnt9b_IRES_EGFP)^pbb48 transgenic embryos with their wild-type heat-shocked siblings (n=4 experiments; *P<0.05; two-tailed, paired Student's t-test). (L) Model depicting how canonical and non-canonical signaling by Wnt9a/b may affect the midline-directed migration of cardiomyocyte progenitor cells, formation of the cardiac cone and leftward jogging. Scale bars: 30 µm.