Department of Pathology, NYU Cancer Institute, New York University School of Medicine, New York, NY 10016, USA.
J Cell Biol. 2013 Jan 21;200(2):141-9. doi: 10.1083/jcb.201209002. Epub 2013 Jan 14.
Proper resolution of stalled replication forks is essential for genome stability. Purification of FBH1, a UvrD DNA helicase, identified a physical interaction with replication protein A (RPA), the major cellular single-stranded DNA (ssDNA)-binding protein complex. Compared with control cells, FBH1-depleted cells responded to replication stress with considerably fewer double-strand breaks (DSBs), a dramatic reduction in the activation of ATM and DNA-PK and phosphorylation of RPA2 and p53, and a significantly increased rate of survival. A minor decrease in ssDNA levels was also observed. All these phenotypes were rescued by wild-type FBH1, but not a FBH1 mutant lacking helicase activity. FBH1 depletion had no effect on other forms of genotoxic stress in which DSBs form by means that do not require ssDNA intermediates. In response to catastrophic genotoxic stress, apoptosis prevents the persistence and propagation of DNA lesions. Our findings show that FBH1 helicase activity is required for the efficient induction of DSBs and apoptosis specifically in response to DNA replication stress.
正确解决停滞的复制叉是基因组稳定性所必需的。FBH1（一种 UvrD DNA 解旋酶）的纯化鉴定了与复制蛋白 A（RPA）的物理相互作用，RPA 是主要的细胞单链 DNA（ssDNA）结合蛋白复合物。与对照细胞相比，FBH1 耗尽的细胞对复制应激的反应表现出明显更少的双链断裂（DSBs），ATM 和 DNA-PK 的激活以及 RPA2 和 p53 的磷酸化显著减少，并且存活率显著增加。还观察到 ssDNA 水平略有下降。所有这些表型都可以通过野生型 FBH1 挽救，但 FBH1 突变体（缺乏解旋酶活性）则不能挽救。FBH1 耗尽对其他形式的遗传毒性应激没有影响，这些形式的 DSBs 形成不需要 ssDNA 中间体。在应对灾难性的遗传毒性应激时，细胞凋亡可防止 DNA 损伤的持续存在和传播。我们的研究结果表明，FBH1 解旋酶活性是 DSBs 和细胞凋亡的有效诱导所必需的，特别是对 DNA 复制应激的反应。
