Ets-1 protects vascular smooth muscle cells from undergoing apoptosis by activating p21WAF1/Cip1 -: Ets-1 regulates basal and inducible p21WAF1/Cip1 transcription via distinct cis-acting elements in the p21WAF1/Cip1 promoter

The cyclin-dependent kinase inhibitor (CKI) p21(WAF1/Cip1) is regulated at the level of transcription by nuclear factors such as the co-activator p300. It is presently unknown whether the Ets family of transcription factors control p21(WAF1/Cip1) gene expression. Ets-1 inhibits apoptosis in vascular smooth muscle cells as determined by both fluorescein isothiocyanate-linked annexin V/propidium iodide staining of cells and fluorescence-activated cell sorting analysis and quantitative cytoplasmic histone-associated internucleosomal DNA fragmentation. p21(WAF1/Cip1) can play a mitogenic and anti-apoptotic role in smooth muscle cells. Using transient transfection and Western blot analysis, we determined that Ets-1 activates p21(WAF1/Cip1) transcription and protein expression. Electrophoretic mobility shift assays revealed that Ets-1 interacts selectively with the (-1350)GGAA(-1347) Ets element in the p21(WAF1/Cip1) promoter. Mutation of this element reduced basal and Ets-1-inducible p21(WAF1/Cip1) promoter-dependent expression. In contrast, the (-1577)GGAT(-1574) motif mediates basal but not Ets-1 activation of the p21(WAF1/Cip1) promoter. Co-immunoprecipitation and co-transfection analysis showed that Ets-1 binds p300 and cooperatively activates p21(WAF1/Cip1) transcription. The phenotypic importance of Ets-1 regulation of p21(WAF1/Cip1) was demonstrated by the capacity of antisense p21(WAF1/Cip1) strategies to block Ets-1-inhibition of apoptosis and inhibit Ets-1-induction of proliferation.