Cytolethal distending toxin of Actinobacillus actinomycetemcomitans induces irreversible cell cycle arrest and cellular distension in various types of mammalian cells. In order to clarify the molecular mechanism of cell cycle arrest induced by CDT, cell cycle regulators in fibroblast and HeLa cells were detected by Western blot analysis with specific antibodies. When treated with CDT, Chk2 activation was observed in both cells. However, in only fibroblasts was accumulation of p53 and p21 observed. In addition, CDT suppressed cyclin B 1, CDC2 and CDK2 in the fibroblasts but not in the HeLa cells. Immunoprecipitation assay with anti-p21 antibody detected CDC2 or CDK2 coupled with p21 in CDT-treated fibroblasts. These results indicated that both CDC2 and CDK2, which represent engines for G2-M and G 1-S progressions respectively, were inactivated by p21 through the p53-dependent mechanism in CDT-treated fibroblasts. Thus, it is suggested that p53 plays an important role in CDT-induced cell cycle arrest in fibroblasts.