Suppression of replication fork progression in low-dose-specic p53-dependent S-phase DNA damage checkpoint
Suppression of replication fork progression in low-dose-specic p53-dependent S-phase DNA damage checkpoint
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DOI：
10.1038/sj.onc.1209624
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发表时间：
2006-09-28
期刊：
影响因子：
8
通讯作者：
Niwa, O.
中科院分区：
文献类型：
--
作者：
Shimura, T.;Toyoshima, M.;Niwa, O.
s期DNA损伤检查点被DNA损伤激活，以延迟DNA合成，从而有时间解决复制障碍。我们之前在辐照精子受精的小鼠受精卵中发现了p53依赖性s期DNA损伤检查点。在这里，我们报道在低剂量照射下，小鼠胚胎成纤维细胞（mef）也存在同样的p53依赖性。p53野生型mef的DNA合成以双相方式受到抑制，在2.5 Gy以下急剧下降，然后更温和地下降至10 Gy。相比之下，p53-/- MEFs在2.5 Gy以下表现出耐辐射DNA合成，而在5 Gy以上细胞保持适度抑制。DNA纤维分析显示，1 Gy辐照抑制p53 WT mef的复制叉进展，但对p53-/- mef无抑制作用。增殖细胞核抗原（PCNA）是DNA聚合酶的钳夹装载者，在1 Gy辐照后在WT MEFs中被磷酸化并重新分布在细胞核内形成病灶。相比之下，在1 gy辐照的p53-/- mef中，PCNA没有磷酸化，也没有与染色质分离。这些结果表明，新的低剂量特异性p53依赖的s期DNA损伤检查点可能通过磷酸化PCNA来调节复制叉的运动。
The S-phase DNA damage checkpoint is activated by DNA damage to delay DNA synthesis allowing time to resolve the replication block. We previously discovered the p53-dependent S-phase DNA damage checkpoint in mouse zygotes fertilized with irradiated sperm. Here, we report that the same p53 dependency holds in mouse embryonic fibroblasts (MEFs) at low doses of irradiation. DNA synthesis in p53 wild-type (WT) MEFs was suppressed in a biphasic manner in which a sharp decrease below 2.5 Gy was followed by a more moderate decrease up to 10 Gy. In contrast, p53-/- MEFs exhibited radioresistant DNA synthesis below 2.5 Gy whereas the cells retained the moderate suppression above 5 Gy. DNA fiber analysis revealed that 1 Gy irradiation suppressed replication fork progression in p53 WT MEFs, but not in p53-/- MEFs. Proliferating cell nuclear antigen (PCNA), clamp loader of DNA polymerase, was phosphorylated in WT MEFs after 1 Gy irradiation and redistributed to form foci in the nuclei. In contrast, PCNA was not phosphorylated and dissociated from chromatin in 1 Gy-irradiated p53-/- MEFs. These results demonstrate that the novel low-dosespecific p53-dependent S-phase DNA damage checkpoint is likely to regulate the replication fork movement through phosphorylation of PCNA.