Phosphorylation of the MADS-Box Transcription Factor MEF2C Enhances Its DNA Binding Activity

doi:10.1074/jbc.271.29.17199

Phosphorylation of the MADS-Box Transcription Factor MEF2C Enhances Its DNA Binding Activity*

From the Department of Molecular Biology and Oncology and the Hamon Center for Basic Cancer Research, The University of Texas, Southwestern Medical Center at Dallas, Dallas, Texas 75235-9148

^§To whom correspondence should be addressed. Tel.: 214-648-1187; Fax: 214-648-1196.

Abstract

Members of the myocyte enhancer factor-2 (MEF2) family of transcription factors activate muscle gene expression by binding an A/T-rich DNA sequence in the control regions of muscle-specific genes. There are four MEF2 factors in vertebrates, MEF2A-D, which share homology in an amino-terminal MADS domain and an adjacent region known as the MEF2 domain, that together mediate DNA binding and dimerization. We show that serine 59 located between the MADS and MEF2 domains of MEF2C is phosphorylated in vivo and can be phosphorylated in vitro by casein kinase-II (CKII). Phosphorylation of this site enhanced the DNA binding and transcriptional activity of MEF2C by increasing its DNA binding activity 5-fold. In vivo ³²P labeling experiments showed that serine 59 is the only phosphorylation site in the MADS and MEF2 domains. Mutagenesis of this serine to an aspartic acid resulted in an increase in DNA binding and transcriptional activity of MEF2C comparable to that observed when this site was phosphorylated, suggesting that phosphorylation augments DNA binding activity by introducing negative charge. This phosphorylation site, which corresponds to a CKII recognition site, is conserved in all known MEF2 factors in organisms ranging from flies to humans, consistent with its importance for the functions of MEF2C.

Footnotes

↵^‡ Supported by an National Institutes of Health postdoctoral fellowship.
↵* This work was supported in part by grants from National Institutes of Health (to E. N. O.), The Human Science Frontiers Program, and The Muscular Dystrophy Association. 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-2

myocyte enhancer factor-2

SRF

serum response factor

PAGE

polyacrylamide gel electrophoresis

EMSA

electrophoretic mobility shift assay

MCK

muscle creatine kinase

MHC

myosin heavy chain

CAT

chloramphenicol acetyltransferase.
↵² M. Kraus, personal communication.
↵³ J. D. Molkentin and E. N. Olson, unpublished results.
↵⁴ J. Zhou and E. N. Olson, unpublished observation.
- Received March 28, 1996.
- Revision received May 1, 1996.