p21Waf1 is required for cellular senescence but not for cell cycle arrest induced by the HDAC inhibitor sodium butyrate

Abstract

Cell senescence is characterized by senescent morphology and permanent loss of proliferative potential. HDAC inhibitors (HDACI) induce senescence and/or apoptosis in many types of tumor cells. Here, we studied the role of cyclin-kinase inhibitor p21waf1 (Cdkn1n gene) in cell cycle arrest, senescence markers (cell hypertrophy, SA-βGal staining and accumulation of γH2AX foci) in p21Waf1+/+ versus p21Waf1-/- mouse embryonic fibroblast cells transformed with E1A and cHa-Ras oncogenes (mERas). While short treatment with the HDACI sodium butyrate (NaB) induced a reversible G1 cell cycle arrest in both parental and p21Waf1-/- cells, long-term treatment led to dramatic changes in p21Waf1+/+ cells only: cell cycle arrest became irreversible and cells become hypertrophic, SA-βGal-positive and accumulated γH2AX foci associated with mTORC1 activation. The p21Waf1+/+ cells lost their ability to migrate into the wound and through a porous membrane. Suppression of migration was accompanied by accumulation of vinculin-staining focal adhesions and Ser3-phosphorylation of cofilin, incapable for F-actin depolymerization. In contrast, the knockout of the p21Waf1 abolished most of the features of NaB-induced senescence, including irreversibility of cell cycle arrest, hypertrophy, additional focal adhesions and block of migration, γH2AX foci accumulation and SA-βGal staining. Rapamycin, a specific inhibitor of mTORC1 kinase, decreased cellular hypertrophy, canceled coffilin phosphorylation and partially restored cell migration in p21Waf1+/+ cells. Taken together, our data indicate a new role of p21Waf1 in cell senescence, which may be connected not only with execution of cell cycle arrest, but also with the development of mTOR-dependent markers of cellular senescence. © 2010 Landes Bioscience.