A Novel Complex Regulates cardiac actin Gene Expression through Interaction of Emb, a Class VI POU Domain Protein, MEF2D, and the Histone Transacetylase p300

Abstract

Expression of the mouse cardiac actin gene depends on a distal enhancer (−7 kbp) which has been shown, in transgenic mice, to direct expression to embryonic skeletal muscle. The presence of this distal sequence is also associated with reproducible expression of cardiac actin transgenes. In differentiated skeletal muscle cells, activity of the enhancer is driven by an E box, binding MyoD family members, and by a 3′ AT-rich sequence which is in the location of a DNase I-hypersensitive site. This sequence does not bind MEF2 proteins, or other known muscle transcription factors, directly. Oct1 and Emb, a class VI POU domain protein, bind to consensus sites on the DNA, and it is the binding of Emb which is important for activity. Emb binds as a major complex with MEF2D and the histone transacetylase p300. The form of Emb present in this complex and as a major form in muscle cell extracts is longer (80 kDa) than that previously described. These results demonstrate the importance of this novel complex in the transcriptional regulation of the cardiac actin gene and suggest a potential role in chromatin remodeling associated with muscle gene activation.

This work was supported by the Pasteur Institute and the C.N.R.S. (URA 1947), with grants from the A.F.M., the M.R.T. (ACI programme), and the E.C. 5th framework programme (Bio4-CT95-0228). S. Molinari received an E.C. Training and Mobility Research fellowship (ERBFMBI-CT95-0240) and an A.F.M. fellowship.

We thank Catherine Bodin for technical assistance, Marc Lavigne for helping with the footprinting analyses, and Marco Pontoglio and Andreas Reinmann for assistance with the ChIP experiments and for helpful comments. We are grateful to R. Prywes, W. Schaffner, and T. Kouzarides for the gift of antibodies and to A. Harel-Bellan for advice on the pulldown assay.