Resolution of R-loops and transcription–replication conflicts by SETX–BRCA1–BARD1 complex

Senataxin (SETX), an RNA–DNA helicase, accumulates at transcription pause sites through the tumor suppressor BRCA1. Here, we provide mechanistic insight into how SETX–BRCA1 resolves transcription-associated R-loops to prevent deleterious outcomes. Specifically, we show that full-length SETX unwinds R-loops with broad specificity and that the complex of BRCA1 and its obligatory partner BARD1 binds R-loops and stimulates R-loop unwinding by SETX. BRCA1–BARD1 alleviates the inhibitory effect of RAD52 on SETX-mediated R-loop unwinding. We also demonstrate that phosphorylation of Ser642 in SETX promotes its interaction with BRCA1 through the tandem BRCT domain of the latter. Accordingly, mutations impacting the catalytic domain or Ser642 in SETX lead to R-loop accumulation, transcription–replication conflicts, replication fork stalling and DNA double-strand breaks in human cells. Thus, our results delineate the molecular basis for functional synergy between SETX and BRCA1–BARD1 in R-loop resolution and the mitigation of transcription–replication conflicts to preserve genome integrity.