Our preliminary studies suggest that Myf5 functions to modify the epigenome in preparation for subsequent transcriptional activation by MyoD, Myogenin and MRF4. Confirming this model will establish a new paradigm for hierarchical families of transcription factors and demonstrate the separation of epigenetic and transcriptional function within members of the same closely related family. Aim 1 will test the hypothesis that the major biological role of Myf5 is to establish active chromatin domains around genes that will be transcribed later in the differentiation program. This aim will extend the preliminary studies and will determine whether Myf5 has predominantly chromatin remodeling activities genome-wide that prepare regions of the genome for transcriptional activation of muscle genes by MyoD and Myogenin. Aim 2 will test the hypothesis that the epigenetic activity of Myf5 prepares genes for subsequent transcriptional activation by Myogenin or MRF4. This aim will determine whether the epigenetic activity of Myf5 facilitates the subsequent DNA binding and transcriptional activity of Myogenin or MRF4 on the expression of the skeletal muscle program. Aim 3 will test the hypothesis that Myf5 has an epigenetic role in mouse muscle development but requires the transcriptional activity of MyoD, Myogenin, or MRF4 for myogenic differentiation of primary mouse myoblasts. Genetic studies have shown that MyoD and Myf5 have largely redundant roles in muscle development and differentiation. This aim will determine whether Myf5 has a distinct epigenetic function in primary mouse muscle cells. Together these studies will provide a new framework for studying the molecular basis of lineage commitment and cell specification in normal development and in cancers.