Abdullah Ummi B, McGouran Joanna F, Brolih Sanja, Ptchelkine Denis, El-Sagheer Afaf H, Brown Tom, McHugh Peter J
Department of Oncology, Weatherall Institute of Molecular Medicine, University of Oxford, Oxford, UK.
Department of Chemistry, University of Oxford, Oxford, UK.
EMBO J. 2017 Jul 14;36(14):2047-2060. doi: 10.15252/embj.201796664. Epub 2017 Jun 12.
During replication-coupled DNA interstrand crosslink (ICL) repair, the XPF-ERCC1 endonuclease is required for the incisions that release, or "unhook", ICLs, but the mechanism of ICL unhooking remains largely unknown. Incisions are triggered when the nascent leading strand of a replication fork strikes the ICL Here, we report that while purified XPF-ERCC1 incises simple ICL-containing model replication fork structures, the presence of a nascent leading strand, modelling the effects of replication arrest, inhibits this activity. Strikingly, the addition of the single-stranded DNA (ssDNA)-binding replication protein A (RPA) selectively restores XPF-ERCC1 endonuclease activity on this structure. The 5'-3' exonuclease SNM1A can load from the XPF-ERCC1-RPA-induced incisions and digest past the crosslink to quantitatively complete the unhooking reaction. We postulate that these collaborative activities of XPF-ERCC1, RPA and SNM1A might explain how ICL unhooking is achieved .
在复制偶联的DNA链间交联（ICL）修复过程中，XPF-ERCC1核酸内切酶对于释放或“解开”ICL的切口是必需的，但ICL解开的机制在很大程度上仍不清楚。当复制叉的新生前导链撞击ICL时，切口被触发。在此，我们报告，虽然纯化的XPF-ERCC1能切割简单的含ICL模型复制叉结构，但模拟复制停滞效应的新生前导链的存在会抑制这种活性。引人注目的是，添加单链DNA（ssDNA）结合复制蛋白A（RPA）可选择性恢复XPF-ERCC1在该结构上的核酸内切酶活性。5'-3'核酸外切酶SNM1A可以从XPF-ERCC1-RPA诱导的切口中加载，并消化穿过交联以定量完成解开反应。我们推测，XPF-ERCC1、RPA和SNM1A的这些协同活性可能解释了ICL是如何解开的。
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