Complex mechanisms underlying impaired activation of Cdk4 and Cdk2 in replicative senescence: Roles of p16, p21, and cyclin D1

Numerous changes in gene expression are known to occur during replicative s enescence, including changes in genes involved in the cell cycle control. I n the present study, we have found a severe impairment in the activation of Cdk2 and Cdk4 in response to mitogens in senescent human fibroblasts and d etermined the molecular basis for this. Although Cdk4 protein was constitut ively expressed in senescent cells at the same level as in early-passage yo ung cells, it was found to be complexed with a distinct set of Cdk inhibito rs, Cdk4 derived from early passage quiescent cells was effectively activat ed by incubation with cyclin D1 and Cdk-activating kinase (CAK) in vitro, w hereas Cdk4 from senescent cells was not. Cdk2 protein was dramatically dec reased in senescent cells and complexed primarily with cyclin D1 and p21. T his cyclin D1-bound Cdk2 was not activated by CAK either in vivo or in vitr o, implicating cyclin D1 as an inhibitor of Cdk2 activation. Thus, one of t he underlying molecular events involved in replicative senescence is the im paired activation of Cdk4 and Cdk2 due to increased binding of p16 to Cdk4 and increased association of Cdk2 with cyclin D1 and p21. (C) 1999 Academic Press.