Orren D K, Petersen L N, Bohr V A
Laboratory of Molecular Genetics, National Institute on Aging, Baltimore, Maryland 21224, USA.
Mol Cell Biol. 1995 Jul;15(7):3722-30. doi: 10.1128/MCB.15.7.3722.
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.
我们研究了紫外线照射对同步化的中国仓鼠卵巢细胞细胞周期进程的影响。通过开发一种使用含羞草碱的新方案实现细胞在S期或G2期的同步化，含羞草碱可在G1/S边界阻断细胞周期进程。去除含羞草碱后，细胞同步进入S期和G2期，从而能够在这两个时期的特定时间点对细胞进行操作。通过血清饥饿一段时间后释放细胞实现细胞在G1期的同步化。通过这些方法同步化的细胞在G1、S和G2期的特定时间点接受紫外线照射，并监测其随后的细胞周期进程。紫外线照射G1期同步化细胞导致进入S期出现剂量依赖性延迟。照射S期同步化细胞会抑制其通过S期的进程，随后导致细胞在G2期长时间积累。在此延长期间注意到一部分细胞发生凋亡。紫外线照射G2期同步化细胞导致G2期阻滞时间较短。阻滞本身及其持续时间分别取决于照射时间（在G2期内）和紫外线剂量。因此，我们定义了一个先前未描述的（在哺乳动物细胞中）G2期紫外线反应检查点。讨论了这些发现对DNA代谢的意义。
