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 poly peptide chains as they emerge from the ribosome. We have identified, c loned, 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 givin g rise to a proline-rich isoform that we termed skNAC. The skNAC prote in was specifically expressed in differentiated myotubes but not in my oblasts. We have identified a specific DNA binding site for skNAC and shown that it can activate transcription through that element. The mur ine myoglobin promoter contains three putative skNAC-binding sites. sk NAC was shown to activate transcription from the myoglobin promoter, a nd site-specific mutation of the skNAC response elements abrogated skN AC-dependent activation. We also examined the role of the NAC isoforms in the myogenic program. Whereas overexpression of alpha NAC prevente d C2C12 differentiation and myotube fusion, the overexpression of skNA C in C2C12 myoblasts led to early fusion of the cells into gigantic my osacs, 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 int o a tissue-specific DNA-binding activator and suggest that this regula tion may be an important event in the proper control of gene expressio n during myogenic differentiation.