TRANSCRIPTIONAL REGULATION OF MUSCLE-SPECIFIC GENES BY ELECTRICAL ACTIVITY

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Skeletal muscle plasticity is dramatically influenced by motoneuron innervation. We have identified muscle genes that are either repressed or stimulated by specific patterns of nerve-derived electrical activity; our goal is to elucidate the molecular mechanisms underlying activity-dependent transcriptional regulation. To this end, we previously demonstrated that myogenin, a muscle-specific factor that regulates transcription of nicotininc acetylcholine receptor (nAChR) genes in cultured cells, is repressed by innervation and de-repressed by denervation. Changes in myogenin levels in muscle preceded receptor regulation during development and after denervation, suggesting that regulation of myogenin may lie upstream of receptor. Analyses of a 3.7 and 1.5 kb myogenin upstream region in transgenic mice have shown that these sequences confer muscle-, developmental-, and denervation-specific regulation. We have developed a modification of a myoblast implantation technique to map transcription regulatory sequences in mature skeletal muscle. Using this technique, we have delineated a 450 bp region that confers the denervation response. To begin mapping the DNA sequences that stimulate the expression of the troponin I slow (TnIs) gene in response to specific patterns of electrical activity, we have begun to map the promoter in cultured C2C12 muscle cells and transgenic mice. Delineation of TnIs upstream sequences in cultured myocytes, demonstrated that 200 bp are necessary and sufficient to confer muscle- and developmental-specific transcription; MyoD and MEF-2 binding sites were shown to be required for activity. However, this region does not direct transcription of the gene in the adult slow muscles of transgenic mice; there is a requirement for sequences further upstream. We have begun to clone and characterize putative trans-acting factors that may regulate myogenin and TnIs transcription. We have isolated cDNAs coding for a novel member of the ets-family and a rat homolog of MEF-2. The functional properties of these factors are currently under investigation. Analysis of the interactions of these families of trans-acting factors should help elucidate the molecular mechanisms underlying fiber-type specific regulation of muscle genes by innervation.