Figure 3

wnt9a is essential for atrioventricular endocardial cell ingression into the cardiac jelly

(A and B) Whole-mount in situ hybridization of wnt9a cardiac expression at 48 hpf. (A) wnt9a expression is mainly within the cardiac tissue of the AVC in wild type (black arrowheads). (B) Upon endothelial-specific overexpression of klf2a (fli1a > klf2a), wnt9a is strongly expressed throughout the entire endocardium.

(C–F) Single confocal z section plane images of the superior AVC endocardium at 72 hpf. (C and D) Normal atrioventricular valvulogenesis with TCF-positive cells on the abluminal side of the forming valve (asterisks) in wild-type (C) and wnt9a^sd49 mutant (D). (E) Endocardial TCF reporter expression is lost upon MO-mediated knockdown of wnt9a. (F) This effect is suppressed in the wnt9a^sd49 mutant background, as confirmed by the presence of TCF-positive cells on the abluminal side of the forming valve (asterisks).

(G and H) Single confocal z section plane images of the AVC endocardium at 96 hpf. (G) In wild type, the primordium of the AV valve leaflet has formed by 96 hpf. (H) Valvulogenesis is completely aborted and the AV endocardium remains monolayered at 96 hpf upon the MO-mediated knockdown of wnt9a.

(I and J) Model depicting the AVC region of the zebrafish endocardium at 96 hpf. (I) Primordium of the wild-type valve leaflet (as in G). (J) AVC endocardium at 96 hpf upon loss of Wnt9a (as in H). Scale bars, 5 μm