Fig. 5

BMP signaling is required for endocardial differentiation.

(A-D) As analyzed by in situ hybridization, myocardial bmp4 expression is greatly reduced in hand2^-/- embryos (B,D) when compared with their siblings at 22hpf (A,B) and 30hpf (C,D). (E-H) Endocardial nfatc1 is greatly downregulated in BMP inhibitor LDN193189-treated embryos at 22hpf (E,F) and 26hpf (G,H) when compared with 0.1% DMSO-treated controls. (I-L) mCherry and GFP fluorescence are observed in the endocardium and myocardium, respectively, in 24 hpf fixed myl7:GFP; kdrl:mCherry embryos that were treated with 10µM LDN193189 from the the tailbud stage onwards (I,K, merged channels; J,L, mCherry channel only). Arrowheads indicate endocardial tube. (M-P) Treatment of embryos with BMP inhibitor LDN193189 starting at the 12- and 17-somite stages results in significant inhibition of nfatc1 expression at 26-28hpf, as analyzed by in situ hybridization. Treatment starting at the 12-somite stage resulted in more significant downregulation of nfatc1 expression than the treatment starting at the 17-somite stage. (Q,R) Myocardial myl7 expression at 30hpf is dysmorphic but does not show significant downregulation in the embryos treated with LDN193189 starting at the tailbud stage. (S,T) Heat-shock-inducible expression of DNBMPR-GFP results in significant downregulation of nfatc1 expression. hsp70:dnBMPR-positive embryos and their siblings were heat-shocked at the tailbud stage for 60min and analyzed at 27hpf.