During embryogenesis, cells from the ventral and dorsal parts of the somites give rise to sclerotome and dermomyotome, respectively. Dermomyotome contains skeletal muscle precursors that are determined by the MyoD family of myogenic factors. We have isolated a novel myogenic repressor, I-mf (Inhibitor of MyoD family), which is highly expressed in the sclerotome. In contrast, MyoD family members are concentrated in the dermomyotome. We demonstrate that I-mf inhibits the transactivation activity of the MyoD family and represses myogenesis. I-mf associates with MyoD family members and retains them in the cytoplasm by masking their nuclear localization signals. I-mf can also interfere with the DNA binding activity of MyoD family members. We postulate that I-mf plays an important role in the patterning of the somite early in development.