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.
Ornatsky
O I
OI
Departments of Kinesiology and Biology, Faculty of Pure and Applied Science, York University, Toronto, Ontario M3J 1P3, Canada.
McDermott
J C
JC
D001704:Biopolymers; D002467:Cell Nucleus / Q000378:metabolism; D004247:DNA / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000235:genetics* / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D026161:MADS Domain Proteins; D064326:MEF2 Transcription Factors; D009132:Muscles / Q000166:cytology / Q000378:metabolism*; D018006:Myogenic Regulatory Factors; D010766:Phosphorylation; D014157:Transcription Factors / Q000235:genetics* / Q000378:metabolism; D014158:Transcription, Genetic
D001704:Biopolymers; D004268:DNA-Binding Proteins; D026161:MADS Domain Proteins; D064326:MEF2 Transcription Factors; C578035:MEF2A protein, human; C578043:MEF2D protein, human; D018006:Myogenic Regulatory Factors; D014157:Transcription Factors; D004247:DNA