MyoD Stimulates RB Promoter Activity via the CREB/p300 Nuclear Transduction Pathway

Abstract

The induction of RB gene transcription by MyoD is a key event in the process of skeletal muscle differentiation, because elevated levels of the retinoblastoma protein are essential for myoblast cell cycle arrest as well as for the terminal differentiation and survival of postmitotic myocytes. We previously showed that MyoD stimulates transcription from the RB promoter independently of direct binding to promoter sequences. Here we demonstrate that stimulation by MyoD requires a cyclic AMP-responsive element (CRE) in the RB promoter, bound by the transcription factor CREB in differentiating myoblasts. We also show that both the CREB protein level and the level of phosphorylation of the CREB protein at Ser-133 rapidly increase at the onset of muscle differentiation and that both remain high throughout the myogenic process. Biochemical and functional evidence indicates that in differentiating myoblasts, MyoD becomes associated with CREB and is targeted to the RB promoter CRE in a complex also containing the p300 transcriptional coactivator. The resulting multiprotein complex stimulates transcription from the RB promoter. These and other observations strongly suggest that MyoD functions by promoting the efficient recruitment of p300 by promoter-bound, phosphorylated CREB.

ACKNOWLEDGMENTS

We are grateful to F. Bex, R. Eckner, R. Goodman, Y. Hamamori, P. Houghton, D. H. Livingston, Y. Nakatani, J. Nyborg, P. L. Puri, V. Sartorelli, and C. Vinson for providing plasmids and critical reagents. We thank L. Pritchard and H. Lehrmann for critical reading of the manuscript. We also thank C. Vesco for helpful discussions during the preparation of the manuscript. We thank Livio Baron, Giancarlo Fontanieri, and Pino Santarelli for expert technical assistance.

This work was supported by grants from Associazione Italiana Ricerca sul Cancro to A.F., from Telethon to M.C., and from European FP5 (grant QLG1-CT-1999-00866) to M.C.