Differential splicing-in of a proline-rich exon converts alpha NAC into a muscle-specific transcription factor

NAC (nascent polypeptide-associated complex) was recently purified as an alpha/beta heterodimeric complex binding the newly synthesized polypeptide chains as they emerge from the ribosome. We have identified, cloned, and characterized a muscle-specific isoform of alpha NAC. The 7.0-kb mRNA arises from differential splicing-in of a 6.0 kb-exon giving rise to a proline-rich isoform that we termed skNAC. The skNAC protein was specifically expressed in differentiated myotubes but not in myoblasts. We have identified a specific DNA binding site for skNAC and shown that it can activate transcription through that element. The murine myoglobin promoter contains three putative skNAC-binding sites. skNAC was shown to activate transcription from the myoglobin promoter, and site-specific mutation of the skNAC response elements abrogated skNAC-dependent activation. We also examined the role of the NAC isoforms in the myogenic program. Whereas overexpression of alpha NAC prevented C2C12 differentiation and myotube fusion, the overexpression of skNAC in C2C12 myoblasts led to early fusion of the cells into gigantic myosacs, suggesting that skNAC may be involved in normal differentiation along the myogenic lineage and in the regulation of myoblast fusion. Our data demonstrate that differential splicing converts alpha NAC into a tissue-specific DNA-binding activator and suggest that this regulation may be an important event in the proper control of gene expression during myogenic differentiation.