PREFERENTIAL BINDING OF MYOD-E12 VERSUS MYOGENIN-E12 TO THE MURINE SARCOMA-VIRUS ENHANCER IN-VITRO

The MyoD family of transcription factors regulates muscle-specific gen e expression in vertebrates. In the adult rat, MyoD mRNA accumulates p redominately in fast-twitch muscle, in particular type IIb and/or IIx fibers, whereas Myogenin mRNA is restricted to slow-twitch type I musc le fibers, Transgenic mice expressing the avian v-ski oncogene from th e murine sarcoma virus (MSV) promoter-enhancer display preferential hy pertrophy of type IIb fast-twitch muscle apparently because of the res tricted expression of the transgene. We tested the hypothesis that pre ferential interactions of MyoD, as a heterodimer with E12, with the MS V enhancer, which has six E-box targets for MyoD family proteins, coul d contribute to v-ski gene expression in IIb muscle fibers, A series o f quantitative binding studies was performed using an electrophoretic mobility shift assay to test MyoD-E12 versus Myogenin-E12 binding to t he MSV enhancer. Our results indicate that MyoD-E12 binds the MSV enha ncer with higher affinity and higher cooperativity than Myogenin-E12. Interestingly, MyoD-E12 bound all of the individual E-boxes tested wit h positive cooperativity indicating DNA-mediated dimerization of the p rotein subunits.