Saur Florian, Lesage Emma, Pradel Lea, Collins Sarah, Finoux Anne-Laure, Alghoul Emile, Le Bozec Benjamin, Rocher Vincent, Carette Romane, Puget Nadine, Couralet Marie, Petiot Melanie, Clouaire Thomas, Marnef Aline, Legube Gaëlle
MCD, Centre de Biologie Intégrative (CBI), CNRS, Université de Toulouse UT, Toulouse, France.
Institut National de la Santé et de la Recherche Médicale (INSERM), Paris, France.
Nat Cell Biol. 2025 May 30. doi: 10.1038/s41556-025-01669-y.
DNA hybrids accumulate at DNA double-strand breaks (DSBs) and were shown to regulate homologous recombination repair. The mechanism responsible for the formation of these non-canonical RNA:DNA structures remains unclear although they were proposed to arise consequently to RNA polymerase II or III loading followed by DSB-induced de novo transcription at the break site. Here, we found no evidence of RNA polymerase recruitment at DSBs. Rather, strand-specific R-loop mapping revealed that RNA:DNA hybrids are mainly generated at DSBs occurring in transcribing loci, from the hybridization of pre-existing RNA to the 3' overhang left by DNA end resection. We further identified the H3K4me3 reader spindlin 1 and the transcriptional regulator PAF1 as factors promoting RNA:DNA hybrid accumulation at DSBs, through their role in mediating transcriptional repression in cis to DSBs. Altogether, we provide evidence that RNA:DNA hybrids accumulate at DSBs occurring in transcribing loci as a result of DSB-induced transcriptional shut down.
DNA杂交体在DNA双链断裂（DSB）处积累，并已证明其可调节同源重组修复。尽管有人提出这些非经典RNA:DNA结构是由于RNA聚合酶II或III加载，随后在断裂位点发生DSB诱导的从头转录而形成的，但负责形成这些结构的机制仍不清楚。在这里，我们没有发现DSB处有RNA聚合酶募集的证据。相反，链特异性R环图谱显示，RNA:DNA杂交体主要在转录位点发生的DSB处产生，是由预先存在的RNA与DNA末端切除留下的3' 突出端杂交形成的。我们进一步确定H3K4me3阅读器纺锤体蛋白1和转录调节因子PAF1是促进RNA:DNA杂交体在DSB处积累的因素，它们通过在顺式调节DSB转录抑制中发挥作用来实现。总之，我们提供的证据表明，由于DSB诱导的转录关闭，RNA:DNA杂交体在转录位点发生的DSB处积累。
