MyoD activity upregulates E2F1 and enhances transcription from the cyclin E promoter in differentiating myoblasts lacking a functional retinoblastoma protein.

We investigated the mechanism leading to cyclin E accumulation when cultured mouse myoblasts, lacking functional Rb because of sequestration or deletion, are exposed to differentiating conditions (mitogen subtraction and cell-cell contact), which activate MyoD and normally downregulate factors involved in cell division. After excluding that stabilization might account for the observed cyclin-E mRNA accumulation, we found an induction of the cyclin-E promoter that correlated with E2F activity upregulation and depended on both MyoD activation and Rb inactivation. Analyses of the E2F1-promoter activity, in normal and Rb-deficient fibroblasts converted by MyoD, identified a MyoD function stimulating E2F1 expression. The E2F1 induction was very manifest in the Rb-/- cells, but also detectable, at the early stage of differentiation, in normal cells. Its effects, although not indispensable for myogenesis, presumably contribute to raise the concentration of Rb-E2F1 transcription-repressing complexes, since MyoD strongly induces also Rb in differentiating myocytes. The activity of an E2F1 promoter lacking the E2F sites indicated that E2F1 itself underwent self-repression by such mechanism at late stages of differentiation. In the absence of Rb, however, the induced E2F1 is left with only its activating role, reversing the normal effect of this MyoD function.