Fig. 7. The Mef2c anterior heart field enhancer is dependent on GATA and ISL sites for function in vivo. The wild-type Mef2c enhancer transgene construct, Mef2c-F6/Frag2-lacZ, or transgenes carrying mutations in either the two GATA sites or the two ISL sites in that context, were used to generate transgenic mice. Representative transgenic embryos at 9.5 days post coitum (dpc) are shown. The wild-type fragment directed strong expression to the pharyngeal mesoderm, outflow tract and right ventricle (A). Mutation of the GATA-p element nearly completely eliminated transgene expression in vivo, although very weak expression could sometimes be seen in the pharyngeal mesoderm. Mutation of the GATA-d site had a dramatic impact on the level of transgene expression, although the pattern of expression remained unchanged. Mutation of both GATA sites in the enhancer (mGATA) completely eliminated lacZ expression (as shown in B). For the double GATA mutant, eight independent transgenic events were analyzed, and none showed any expression in the heart. Mutation of both ISL sites (mISL) in the enhancer completely eliminated all transgene expression (C) in all six independent transgenic events analyzed. The wild-type 3970 bp enhancer construct directed robust lacZ expression, as shown in (A), in 16 of 17 transgenic lines. Arrowheads denote expression in the pharyngeal mesoderm/anterior heart field. RV, right ventricle.