Fig. 5. Proximal, high-affinity GATA sites are critical for expression of the XMLC2 promoter. Heart-specific expression is unaffected by targeted mutation of individual GATA sites, such as GATA#2 (A). When both GATA#2 and GATA#3 sites are inactivated, expression in the heart is maintained, but it is weak (B; ventral view). Mutation of both GATA#1 and GATA#2 abolishes activity of the promoter (C). Note that transgenesis in this case is confirmed by expression of the co-transgene gamma-crystallin in the eye and hindbrain. (D) GATA#1 and GATA#2 have greater affinity for GATA4 than the more distal GATA#3. S, specific competitor (unlabelled probe); N, non-specific competitor (both used at 100x molar excess). (E) Competition assay, using 20, 50 and 100x molar excess of competitors confirms this result.