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Fig. 9
FGF signaling functions upstream of Nkx genes to repress amhc expression in the ventricle. (A-D) In situ hybridization showing nkx2.5 (A,B) and nkx2.7 (C,D) expression in dorsal views at 26.5?hpf, following heat shock at 18?hpf. Nontransgenic embryos (A,C) exhibit robust expression of nkx2.5 (n=24) and nkx2.7 (n=12), and Tg(hsp70:dnfgfr1) embryos (B,D) exhibit reduced expression of nkx2.5 (n=24) and nkx2.7 (n=12). This reflects a reduced amount of ventricular tissue in Tg(hsp70:dnfgfr1) embryos (compare with Fig. 4M), as well as lower expression levels within this tissue. (E-G) In situ hybridization showing amhc expression at 48?hpf in nontransgenic (F) and Tg(hsp70:nkx2.5) (E,G) embryos treated with DMSO (E) or SU5402 (F,G); frontal views. In some cases (G), overexpression of nkx2.5 represses ectopic amhc expression in Tg(hsp70:nkx2.5) embryos that were treated with SU5402 at 18?hpf and subsequently heat shocked at 24?hpf. (H) Table reports the results of experiments gauging whether amhc expression in the ventricle of SU5402-treated embryos can be repressed by overexpression of nkx2.5. Asterisk indicates a statistically significant difference in the frequency of detecting ectopic amhc expression, compared to nontransgenic siblings (P=0.0019, Fisher?s exact test), representing partial rescue of the SU5402-treated phenotype. The degree of rescue was not enhanced when we used embryos carrying two copies of Tg(hsp70:nkx2.5) or when we administered two rounds of heat shock (data not shown). Scale bars: 50??m.