A UV-responsive G₂ checkpoint in rodent cells

We have studied the effect of UV irradiation on the cell cycle progression of synchronized Chinese hamster ovary cells. Synchronization of cells in S or G₂ phase was accomplished by the development of a novel protocol using mimosine, which blocks cell cycle progression at the G₁/S boundary. After removal of mimosine, cells proceed synchronously through the S and G₂ phases, allowing manipulation of cells at specific points in either phase. Synchronization of cells in G₁ was achieved by release of cells after a period of serum starvation. Cells synchronized by these methods were UV irradiated at defined points in G₁, S, and G₂, and their subsequent progression through the cell cycle was monitored. UV irradiation of G₁- synchronized cells caused a dose-dependent delay in entry into S phase. Irradiation of S-phase-synchronized cells inhibited progression through S phase and then resulted in accumulation of cells for a prolonged interval in G₂. Apoptosis of a subpopulation of cells during this extended period was noted. UV irradiation of G₂-synchronized cells caused a shorter G₂ arrest. The arrest itself and its duration were dependent upon the timing (within G₂ phase) of the irradiation and the UV dose, respectively. We have thus defined a previously undescribed (in mammalian cells) UV-responsive checkpoint in G₂ phase. The implications of these findings with respect to DNA metabolism are discussed.