Overexpression of cyclin D1, a G1 cell cycle regulator, is often found in many different tumor types, such as breast carcinoma and squamous cell carcinoma of the head and neck. The overexpression of this protein is, in several cases, associated with a poor prognosis. In this study, the effect of cyclin D1 on radiosensitivity was investigated in a breast tumor cell line, MCF7, containing a cyclin D1 gene construct under the control of a tetracycline-sensitive regulator. MCF7 cells cultured without tetracycline resulted in a 6-fold increase in the cyclin D1 protein. Cyclin D1-overexpressing MCF7 cells were more sensitive to ionizing radiation than the nonoverexpressing counterparts. The cyclin D1-overexpressing cells also exhibited a higher induction of apoptosis. Treatment with a dose of 5 Gy resulted in a rapid increase of p53 and p21 in the cyclin D1-overexpressing cells. Nonoverexpressing cells showed a more transient expression of these proteins after ionizing radiation. A pronounced G2-M block was observed in both cell lines. The cyclin D1-overexpressing cells were, however, released earlier from the block than the control cells. These data suggest that overexpression of cyclin D1 alters sensitivity toward ionizing radiation by modulating gamma-radiation-induced G2-M transition.