CDC45 and DPB11 are required for processive DNA replication and resistance to DNA topoisomerase I-mediated DNA damage
CDC45 and DPB11 are required for processive DNA replication and resistance to DNA topoisomerase I-mediated DNA damage
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DOI：
10.1073/pnas.96.20.11440
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发表时间：
1999-09-28
影响因子：
11.1
通讯作者：
Bjornsti, MA
中科院分区：
文献类型：
--
作者：
Reid, RJD;Fiorani, P;Bjornsti, MA
抗肿瘤药物喜树碱通过可逆地稳定共价酶-DNA中间体来靶向DNA拓扑异构酶I。DNA复制叉随后与这些药物-酶-DNA复合体碰撞，产生细胞毒性DNA损伤，发出细胞周期停滞的信号，最终导致细胞死亡。尽管进行了密集的调查，但所产生的损害的特征和解决损害的修复过程仍然不清楚。酵母遗传筛选用于分离对DNA拓扑异构酶I介导的DNA损伤具有更高敏感性的条件突变株。选择了在DNA拓扑异构酶I突变体Top1T722Ap存在下表现出温度敏感型生长的细胞。Ala取代Thr722增加了共价Top1T722Ap-DNA中间体的稳定性，模拟了喜树碱的细胞毒作用。分离到的两个突变体，cdc45-10和dpb11-10，在没有DNA损伤剂的情况下，表现出DNA复制的特殊缺陷和合成的致死表型。在不允许的条件下积累冈崎片段表明，通过聚合酶交换促进DNA复制是一种常见的功能。这些结果为理解喜树碱和DNA拓扑异构酶I诱导的DNA损伤的细胞过程提供了机制基础。
The antitumor agent camptothecin targets DNA topoisomerase I by reversibly stabilizing a covalent enzyme-DNA intermediate. The subsequent collision of DNA replication forks with these drug-enzyme-DNA complexes produces the cytotoxic DNA lesions that signal cell cycle arrest and ultimately lead to cell death. Despite intense investigation, the character of the lesions produced and the repair processes that resolve the damage remain poorly defined. A yeast genetic screen was implemented to isolate conditional mutants with enhanced sensitivity to DNA topoisomerase I-mediated DNA damage. Cells exhibiting temperature-sensitive growth in the presence of the DNA topoisomerase I mutant, Top1T722Ap, were selected. Substitution of Ala for Thr722 increases the stability of the covalent Top1T722Ap-DNA intermediate, mimicking the cytotoxic action of camptothecin. Two mutants isolated, cdc45-10 and dpb11-10, exhibited specific defects in DNA replication and a synthetic lethal phenotype in the absence of DNA damaging agents. The accumulation of Okazaki fragments under nonpermissive conditions suggests a common function in promoting processive DNA replication through polymerase switching. These results provide a mechanistic basis for understanding the cellular processes involved in the resolution of DNA damage induced by camptothecin and DNA topoisomerase I.