The incidence of DNA mutation and subsequent risk of transformation in different cell types may depend on cell type-specific variation in position and duration of cell cycle arrest after exposure to DNA-damaging agents. To determine whether cell type-specific checkpoints occur, normal human epidermal keratinocytes (HKs) and human dermal fibroblasts (HFs), isolated from the same tissue, were exposed to genotoxic agents. Following exposure, cell cycle arrest profiles, cell proliferation rates, and select protein levels and activities were analyzed and found to be cell type dependent. After exposure to either gamma-radiation or Adriamycin, HFs arrested primarily in G1, whereas HKs arrested predominantly in G2. The attenuated G1 arrest in the HKs correlated with less p53 protein accumulation, as compared to that observed in G1-arrested HFs. Although gamma-irradiated HFs were unable to reenter the cell cycle, HKs began proliferating 72 h posttreatment. Consistent with the cell cycle profiles observed, cyclin-dependent kinase activities were inhibited for a longer duration in HFs as compared to HKs after gamma-irradiation. The results indicate that cell cycle checkpoint response to genotoxic insult may vary according to cell type within any given tissue. The attenuated G1 arrest observed in HKs may be an important factor in the transforming events leading to skin neoplasia.