MEF2 protein expression, DNA binding specificity and complex composition, and transcriptional activity in muscle and non-muscle cells

Tissue-specific gene expression can be mediated by complex transcriptional regulatory mechanisms. Based on the dichotomy of the ubiquitous distribution of the myocyte enhancer factor 2 (MEF2) gene mRNAs compared to their cell type-restricted activity, we investigated the basis for their tissue specificity. Electrophoretic mobility shift assays using the muscle creatine kinase MEF2 DNA binding site as a probe showed that HeLa, Schneider, L6E9 muscle, and C2C12 muscle cells have a functional MEF2 binding activity that is indistinguishable based on competition analysis. Interestingly, chloramphenicol acetyltransferase reporter assays showed MEF2 site-dependent trans-activation in myogenic C2C12 cells but no trans-activation by the endogenous MEF2 proteins in HeLa cells. By immunofluorescence, we detected abundant nuclear localized MEF2A and MEF2D protein expression in HeLa cells and C2C12 muscle cells. Using immuno-gel shift analysis and also co-immunoprecipitation studies, we show that the predominant MEF2 DNA binding complex bound to MEF2 sites from either the muscle creatine kinase or c-jun regulatory regions in C2C12 muscle cells is comprised of a MEF2A homodimer, whereas in HeLa cells, it is a MEF2A:MEF2D heterodimer. Thus, the presence of MEF2 DNA binding complexes is not necessarily coupled with trans-activation of target genes. The ability of the MEF2 proteins to activate transcription in vivo correlates with the specific dimer composition of the DNA binding complex and the cellular context.