Liver cells from p21Cip1−/− mice subjected to partial hepatectomy (PH) progress into DNA synthesis faster than those from wild-type mice. These cells also show a premature induction of cyclin E/cyclin-dependent kinase (CDK) 2 activity. We studied the mechanisms whereby cells lacking p21Cip1 showed a premature induction of this activity. Whereas the levels of CDK2, cyclin E, and p27Kip1 were similar in both wild-type and p21Cip1−/− mice, those of the activator CDC25A were much higher in p21Cip1−/− quiescent and regenerating livers than in wild-type animals. Moreover, p21Cip1−/− cells also showed a premature translocation of CDC25A from cytoplasm into the nucleus. The ectopic expression of p21Cip1 into mice embryo fibroblasts from p21Cip1−/− mice decreased the levels of CDC25A and delayed its nuclear translocation. The levels of CDC25A messenger RNA in p21Cip1−/− cells were higher than in wild-type cells, suggesting that this increase might be responsible, at least in part, for the high levels of CDC25A protein in these cells. Thus, the results reported here indicate that p21Cip1 regulates the levels and the intracellular localization of CDC25A. We also found a good correlation between CDC25A nuclear translocation and cyclin E/CDK2 activation. In conclusion, premature translocation of CDC25A to the nucleus might be involved in the advanced induction of cyclin E/CDK2 activity and DNA replication in cells from animals lacking p21Cip1.