Fig. 1. Endocardial-restricted inactivation of Gata4. (A) Structure of wild-type (wt), hypomorphic (H), floxed, and deleted (ex2) Gata4 alleles. (B) Fate mapping of T2Cre-recombined endothelial cells. (B1,B2) In T2Cre+; R26RstoplacZ embryos at E9.5, ß-galactosidase reporter expression (blue) was seen in the endocardium (arrow) and EC mesenchyme (arrowhead) of the AVC, and in the OT endothelium. (B3) By E11.5, the ß-galactosidase-positive cells populated the AV cushions (asterisks). Endothelial-derived OT cushion mesenchyme (yellow arrow) was largely confined to the most proximal portion of the OT cushion. Most of the outflow cushion was not recombined by T2Cre (green arrow). (C) Endocardial-restricted inactivation of Gata4 by T2Cre at E9.5. Control (C1,C2) and Gata4^T2del (C3,C4) embryos, hybridized to a Gata4 exon 2-specific probe (red pseudocolor). Blue, DAPI counterstain. The Gata4 exon 2 in situ hybridization signal was present in the endocardium of control embryos but not of mutant embryos (white arrowheads). Gata4 expression in the myocardium (yellow arrowheads) and proepicardium (star) was unaffected. (D) Inactivation of Gata4 in endothelium and endothelium-derived cells at E11.5. In control embryos (D1), Gata4 exon 2 in situ hybridization signal was present in the endocardium (white arrowhead), myocardium (yellow arrowhead), and AV and OT cushion mesenchyme. In mutant embryos (D2), Gata4 was not detected in endothelium (white arrowhead) or AVC mesenchyme. Strong signal was still present in mid and distal OT cushion mesenchyme (green arrow) and myocardium (yellow arrowhead). a, atria; v, ventricle; ot, OT. Scale bars in C and D: 100 µm.