MEF2 Protein Expression, DNA Binding Specificity and Complex Composition, and Transcriptional Activity in Muscle and Non-muscle Cells

doi:10.1074/jbc.271.40.24927

MEF2 Protein Expression, DNA Binding Specificity and Complex Composition, and Transcriptional Activity in Muscle and Non-muscle Cells*

From the Departments of Kinesiology and Biology, Faculty of Pure and Applied Science, York University, Toronto, Ontario M3J 1P3, Canada

^‡ To whom correspondence should be addressed:
341 Farquharson LSB, York University, 4700 Keele St., Toronto, Ontario, M3J 1P3 Canada
. Tel.: 416-736-2100, ext. 30389; Fax: 416-736-5698; E-mail: FS300557{at}sol.yorku.ca

Abstract

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.

Footnotes

↵* This work was supported by a grant from the Natural Sciences and Engineering Research Council of Canada. The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
↵¹ The abbreviations used are:

MEF

myocyte enhancer factor

CAT

chloramphenicol acetyltransferase

MCK

muscle creatine kinase

CIP

calf intestinal phosphatase

TK

thymidine kinase.
↵² R. Prywes, personal communication.
↵³ O. Ornatsky and J. McDermott, unpublished observation.
- Received June 13, 1996.