DNA Replication Group, Molecular Oncology Programme, Spanish National Cancer Research Centre (CNIO), Melchor Fernández Almagro 3, 28029, Madrid, Spain.
Institute of Molecular Cancer Research, University of Zurich, Winterthurerstrasse 190, 8057, Zurich, Switzerland.
EMBO J. 2024 Mar;43(6):1043-1064. doi: 10.1038/s44318-024-00038-z. Epub 2024 Feb 15.
Eukaryotic cells rely on several mechanisms to ensure that the genome is duplicated precisely once in each cell division cycle, preventing DNA over-replication and genomic instability. Most of these mechanisms limit the activity of origin licensing proteins to prevent the reactivation of origins that have already been used. Here, we have investigated whether additional controls restrict the extension of re-replicated DNA in the event of origin re-activation. In a genetic screening in cells forced to re-activate origins, we found that re-replication is limited by RAD51 and enhanced by FBH1, a RAD51 antagonist. In the presence of chromatin-bound RAD51, forks stemming from re-fired origins are slowed down, leading to frequent events of fork reversal. Eventual re-initiation of DNA synthesis mediated by PRIMPOL creates ssDNA gaps that facilitate the partial elimination of re-duplicated DNA by MRE11 exonuclease. In the absence of RAD51, these controls are abrogated and re-replication forks progress much longer than in normal conditions. Our study uncovers a safeguard mechanism to protect genome stability in the event of origin reactivation.
真核细胞依赖几种机制来确保基因组在每个细胞分裂周期中准确复制一次，防止 DNA 过度复制和基因组不稳定。这些机制中的大多数限制了起始许可蛋白的活性，以防止已经使用过的起始点的重新激活。在这里，我们研究了在起始点重新激活的情况下，是否有其他控制来限制重新复制的 DNA 的延伸。在强迫起始点重新激活的细胞的遗传筛选中，我们发现重新复制受到 RAD51 和 FBH1 的限制，FBH1 是 RAD51 的拮抗剂。在结合在染色质上的 RAD51 的存在下，源自重新激活的起始点的叉被减缓，导致叉反转的频繁发生。最终由 PRIMPOL 介导的 DNA 合成的重新起始会产生 ssDNA 缺口，这有助于 MRE11 核酸外切酶部分消除重新复制的 DNA。在没有 RAD51 的情况下，这些控制被废除，重新复制叉的进展比正常情况下长得多。我们的研究揭示了一种保护机制，可以在起始点重新激活时保护基因组稳定性。
