Fig. 7. Binding of MyoD and MEF2 to the Mef2c skeletal muscle regulatory region. (A) Sequence of the minimal skeletal muscle regulatory region is shown. The MEF2-binding site extends from nucleotides -64 to -56. The E-box includes nucleotides -38 through -33. (B) Schematic representation of the 1.1 kb Mef2c skeletal muscle control region. The region encompassing nucleotides -512 to +41, with respect to the transcriptional start site of Mef2c (see Fig. 1). The specific MEF2 site and E-box nucleotides mutated for the experiments shown in Fig. 8 are indicated beneath the wild-type sequence. -³²P-labeled oligonucleotides for the (C) MEF2 site and (D) E-box of the Mef2c skeletal muscle regulatory region were used as probes in gel mobility shift assays with the in vitro translated (C) Myc-tagged MEF2C or (D) MyoD and E12 proteins. Gel shift reactions using unprogrammed reticulocyte lysate are included in parallel lanes. A 100-fold excess of unlabeled (C) MEF2 site or (D) E-box oligonucleotides was used as competitor. (C) Anti-Myc and (D) anti-MyoD antibodies were used for supershifts. Labeled arrows indicate the positions of the various protein-DNA complexes.