A large body of evidence has demonstrated that normal human fibroblasts have a limited division potential in culture and underwent senescence, a process whereby cells became arrested in the G1 phase of the cell cycle and overexpressed a DNA synthesis inhibitor(s). Cyclin-dependent kinase two (Cdk2) is required for the promotion of the Gi-to-S phase transition in human cells. Senescent fibroblasts contain intact cyclin-Cdk2 complexes but cannot induce Cdk2 protein kinase activity in response to mitogen stimulation. Recently, we cloned p21Sdi1, a potent inhibitor of DNA synthesis and Cdk2 kinase activity, from a senescent cell cDNA library and demonstrated that it was expressed at significantly higher levels in senescent cells than actively proliferating cells. In contrast to actively dividing cells, mitogen-stimulated senescent cells do not down-regulate the expression of p21Sdi1 and do not express late G1 phase gene products that are required for entry into S phase. We suggest that the inability of mitogen-stimulated senescent cells to down-regulate p21Sdi1 levels contributes to the resulting lack of late Gi gene expression and failure to traverse the G1/S phase boundary.