期刊
CELL CYCLE
卷 5, 期 16, 页码 1823-1830出版社
TAYLOR & FRANCIS INC
DOI: 10.4161/cc.5.16.3009
关键词
G1/S; cell cycle checkpoints; DNA damage; DNA double-strand breaks; ionizing radiation; replication origin regulation
类别
DNA double-strand breaks caused by ionizing radiation have been shown to induce G(1)/S-i intra-S-phase, and G(2)/M cell cycle checkpoints. However, analysis of the immediate induction of G(1)/S checkpoint at a cellular level has been hampered by the inability to distinguish cells that were already replicating DNA at the time of damage from cells that entered S phase following the DNA damage. We have developed a novel strategy for assessing the initiation of the G(1)/S checkpoint following g-irradiation within asynchronous, low passage, primary mouse embryonic fibroblast cultures (MEFs) using a staggered CldU/IdU double-labeling protocol. Contrary to the current model of the G(1)/S checkpoint, we found that 65% of late-G1 primary MEFs still proceeded into S phase after a gamma-irradiation dose of 5 Gy. The delayed p53-dependent G(1)/S checkpoint was intact in these cells, and a G(2)/M checkpoint that was over 90% effective was induced within one hour and maintained through six hours post-irradiation. Furthermore, these cells also exhibited an intra-S-phase replication slow-down, as there was a decrease in the S/G(2) transition frequency of primary MEFs following gamma-irradiation. The absence of an immediate G(1)/S checkpoint in primary MEFs suggests that in late G(1) these cells may predominantly respond to DNA damage at the level of individual replication origins, rather than by inducing a complete shut-down of S-phase entry.
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