p21Waf1/Cip1/Sdi1 induces permanent growth arrest with markers of replicative senescence in human tumor cells lacking functional p53.

We have shown previously that wild type p53 can rapidly induce replicative senescence in EJ human bladder carcinoma cells lacking functional p53. A major effector of p53 functions is p21Waf1/Cip1/Sdi1, a potent cyclin-dependent kinase inhibitor. p21Waf1/Cip1/Sdi1 has been shown to be involved in both p53 dependent and independent control of cell proliferation, differentiation and death. To directly investigate the effects of p21Waf1/Cip1/Sdi1 in the p53 response observed in EJ tumor cells, we established p21Waf1/Cip1/Sdi1 inducible lines using the tetracycline-regulatable vector system. p21Waf1/Cip1/Sdi1 induction caused irreversible cell cycle arrest in both G1 and G2/M, and diminished Cdk2 kinase activity. In addition, p21Waf1/Cip1/Sdi1 induction led to morphological alterations characteristic of cells undergoing replicative senescence with morphological, biochemical and ultrastructural markers of the senescent phenotype. Furthermore, sustained p21Waf1/Cip1/Sdi1 induction sensitized EJ cells to apoptotic cell death induced by mitomycin C, a cross-linking DNA damaging agent. These findings support the function of p21Waf1/Cip1/Sdi1 as an inducer of replicative senescence and a major mediator of this phenomenon in response to p53. Moreover, our results imply that therapeutic intervention in human cancers might be aimed at sustained elevation of p21Waf1/Cip1/Sdi1 expression.