The Myogenic Regulatory Factor (MRF) family is composed of 4 members:  Myf-3 (MyoD), Myf-4 (myogenin), Myf-5 and Myf-6 (MRF4/herculin). These proteins are involved in the skeletal muscle differentiation program that balances myoblast proliferation vs. differentiation. Activation of this program in response to growth factor withdrawal is accompanied by myoblast cell fusion to form multinucleated myotubes, irreversible commitment of muscle cells to the post-mitotic state, and transcriptional activation of muscle-specific genes. MyoD is expressed in undifferentiated myogenic proliferating cells. Expression of MyoD family genes activates myogenesis in various non-muscle cell types. In mice bearing a null Myf-5/MyoD double mutation, muscle fibers and myoblasts are absent. MyoD acts downstream of Pax-3 and Myf-5 in skeletal myogenesis and is sensitive to growth factors and transforming gene products such as Ras and Fos, which have been shown to abolish the expression of MyoD in myoblasts. MyoD induces muscle-specific alternative splicing of mouse mitochondrial ATP synthase gamma-subunit, beta-tropomyosin, MEF2A and MEF2D pre-mRNAs.

MyoD is a transcription factor with a basic helix-loop-helix (bHLH) DNA-binding domain that mediates dimerization and binding to a DNA consensus sequence termed an E-box (CANNTG). These E-boxes are present in the control regions of several skeletal muscle-specific genes such as the muscle creatinine kinase, myosin light chains 1 and 3, desmin and the acetylcholine receptor. Its dimeric partners are E-box binding proteins such as Tal-1, E12 and E476. While MyoD and E47 both form homodimeric complexes with DNA that have similar dissociation constants, a stochiometric mixture of these bHLH domains forms almost exclusively heterodimeric complexes on DNA. The DNA binding activity of MyoD is repressed by forming heteromeric complexes with inhibitory proteins, such as Id protein and Mist1 transcription factor, which occupy the specific E-box target sites.