ARTICLE https://doi.org/10.1038/s41467-021-24201-w OPEN UPF1 promotes the formation of R loops to stimulate DNA double-strand break repair ✉ Greg H. P. Ngo 1, Julia W. Grimstead1 & Duncan M. Baird 1 DNA-RNA hybrid structures have been detected at the vicinity of DNA double-strand breaks (DSBs) occurring within transcriptional active regions of the genome. The induction of DNA- RNA hybrids strongly affects the repair of these DSBs, but the nature of these structures and 1234567890():,; how they are formed remain poorly understood. Here we provide evidence that R loops, three-stranded structures containing DNA-RNA hybrids and the displaced single-stranded DNA (ssDNA) can form at sub-telomeric DSBs. These R loops are generated independently of DNA resection but are induced alongside two-stranded DNA-RNA hybrids that form on ssDNA generated by DNA resection. We further identified UPF1, an RNA/DNA helicase, as a crucial factor that drives the formation of these R loops and DNA-RNA hybrids to stimulate DNA resection, homologous recombination, microhomology-mediated end joining and DNA damage checkpoint activation. Our data show that R loops and DNA-RNA hybrids are actively generated at DSBs to facilitate DNA repair. 1 Division of Cancer and Genetics, School of Medicine, Cardiff University, Cardiff, UK. ✉email:
[email protected] NATURE COMMUNICATIONS | (2021) 12:3849 | https://doi.org/10.1038/s41467-021-24201-w | www.nature.com/naturecommunications 1 ARTICLE NATURE COMMUNICATIONS | https://doi.org/10.1038/s41467-021-24201-w elomeres are nucleoprotein structures at the end of linear Results Tchromosomes that protect the DNA from being recognized DNA resection contributes to deletion formation at telomere as DNA double-strand breaks (DSBs).