The skeletal muscle regulatory factors (MRFs), of which myoD is the prototype, are a family of helix-loop-helix proteins that act as transcription factors for skeletal muscle-specific gene expression. The various members of the MRF family are differentially expressed during development suggesting that they may participate in orchestrating developmental changes in skeletal muscle maturation. To address this, we have focussed on the changes in MRF gene expression that occur between embryonic and fetal stages of chicken skeletal muscle development. This transition involves the switching of contractile protein isoform gene expression from the """"""""cardiac"""""""" to the """"""""skeletal"""""""" type, occurs in concert with increases in muscle activity, and is associated with increased protein kinase C (PKC) activity. In work completed this year, we show that expression of the MRFs and """"""""cardiac"""""""" type contractile proteins mRNA in cultured muscle cells are decreased when PKC levels are elevated either pharmacologically or genetically. We also show that the effect of PKC on myoD is at the level of gene transcription, but cannot be overcome by overexpression of myoD driven by a heterologous promoter. Since MRFs are likely to positively regulate their own gene expression, the data is consistent with a model of regulation in which the primary effect of PKC is post-transcriptional, creating a defunct transcription factor that can neither transactivate its own gene or that of its targets. Additional information suggests that the effect is likely mediated by direct phosphorylation of a threonine residue in the DNA binding domain of the MRFs.
