Department of Biology, University of Iowa, Iowa City, IA, USA.
Department of Molecular and Human Genetics, Baylor College of Medicine, Houston, TX, USA.
Nature. 2021 Feb;590(7847):655-659. doi: 10.1038/s41586-020-03172-w. Epub 2021 Jan 20.
Break-induced replication (BIR) repairs one-ended double-strand breaks in DNA similar to those formed by replication collapse or telomere erosion, and it has been implicated in the initiation of genome instability in cancer and other human diseases. Previous studies have defined the enzymes that are required for BIR; however, understanding of initial and extended BIR synthesis, and of how the migrating D-loop proceeds through known replication roadblocks, has been precluded by technical limitations. Here we use a newly developed assay to show that BIR synthesis initiates soon after strand invasion and proceeds more slowly than S-phase replication. Without primase, leading strand synthesis is initiated efficiently, but is unable to proceed beyond 30 kilobases, suggesting that primase is needed for stabilization of the nascent leading strand. DNA synthesis can initiate in the absence of Pif1 or Pol32, but does not proceed efficiently. Interstitial telomeric DNA disrupts and terminates BIR progression, and BIR initiation is suppressed by transcription proportionally to the transcription level. Collisions between BIR and transcription lead to mutagenesis and chromosome rearrangements at levels that exceed instabilities induced by transcription during normal replication. Together, these results provide fundamental insights into the mechanism of BIR and how BIR contributes to genome instability.
断裂诱导复制 (BIR) 修复 DNA 中的单端双链断裂，类似于复制崩溃或端粒侵蚀形成的断裂，并且它与癌症和其他人类疾病中基因组不稳定性的起始有关。先前的研究已经定义了 BIR 所需的酶；然而，由于技术限制，对初始和扩展的 BIR 合成以及迁移的 D 环如何通过已知的复制障碍的理解受到了阻碍。在这里，我们使用新开发的测定法表明，BIR 合成在链入侵后很快开始，并比 S 期复制进行得更慢。没有引发酶，前导链合成可以有效地起始，但无法进行超过 30 千碱基，这表明引发酶对于稳定新生前导链是必需的。可以在没有 Pif1 或 Pol32 的情况下起始 DNA 合成，但不能有效地进行。间质端粒 DNA 会破坏和终止 BIR 进程，并且转录与转录水平成比例地抑制 BIR 起始。BIR 与转录之间的碰撞导致诱变和染色体重排的水平超过正常复制过程中转录引起的不稳定性。总之，这些结果为 BIR 的机制以及 BIR 如何导致基因组不稳定性提供了基本的见解。
