Regulation of p 21^WAF1/CIP1 Expression during Cellular Differentiation

Abstract

In the present study, the molecular mechanisms regulating p 21WAF1/Cipl expression by basic helix-loop-helix (bHLH) factors during cellular differentiation were investigated. The cyclin dependent kinase inhibitor p 21 plays crucial roles during differentiation of osteoblasts and myoblasts. In the osteoblastic cell line MG 63, expression of the p 21 gene has been shown to be upregulated by E 2 A factors, members of the bHLH factor family. In addition, E 2 A-dependent activation of p 21 promoter can be inhibited by another bHLH factor, TWIST. Using reporter assays with mutant p 21 promoters, a novel element was identified in the p 21 promoter, which is essential for E 2 A-dependent activation and TWIST-mediated inhibition. Interestingly, in the myoblastic cell line C 2 C 12, this sequence was not involved in E 2 A-dependent activation of p 21 expression. Gel mobility shift assays showed a specific complex of the novel p 21 promoter element with nuclear factor (s) of MG 63 cells. Complex formation was inhibited by the addition of anti-TWIST antibody. In contrast no complexes could be identified with C 2 C 12 cells. These results raise the possibility that interactions of the bHLH factors with the novel p 21 promoter element are cell type specific. This suggests a novel mechanism regulating p 21 expression.