BRCA2 promotes DNA‐RNA hybrid resolution by DDX5 helicase at DNA breaks to facilitate their repair‡

Abstract

The BRCA2 tumor suppressor is a DNA double-strand break (DSB) repair factor essential for maintaining genome integrity. BRCA2deficient cells spontaneously accumulate DNA-RNA hybrids, a known source of genome instability. However, the specific role of BRCA2 on these structures remains poorly understood. Here we identified the DEAD-box RNA helicase DDX5 as a BRCA2-interacting protein. DDX5 associates with DNA-RNA hybrids that form in the vicinity of DSBs, and this association is enhanced by BRCA2. Notably, BRCA2 stimulates the DNA-RNA hybrid-unwinding activity of DDX5 helicase. An impaired BRCA2-DDX5 interaction, as observed in cells expressing the breast cancer variant BRCA2-T207A, reduces the association of DDX5 with DNA-RNA hybrids, decreases the number of RPA foci, and alters the kinetics of appearance of RAD51 foci upon irradiation. Our findings are consistent with DNA-RNA hybrids constituting an impediment for the repair of DSBs by homologous recombination and reveal BRCA2 and DDX5 as active players in their removal.

New Faces of old Friends: Emerging new Roles of RNA-Binding Proteins in the DNA Double-Strand Break Response

“…Interestingly, upon DSB recruitment, CIRBP is poly (ADPribosyl)ated and then rapidly excluded again from the damaged chromatin (Chen et al, 2018). Similar association and dissociation dynamics have been observed for other RBPs as well (Salton et al, 2010;Ha et al, 2011;Adamson et al, 2012;Beli et al, 2012;Polo et al, 2012;Britton et al, 2014;Altmeyer et al, 2015;Yu et al, 2020;Sessa et al, 2021). Another RBP that is recruited to damaged chromatin upon DSB formation and that appears to directly regulate DSB signaling is WRAP53 (WD40encoding RNA antisense to p53, also known as TCAB1).…”

“…As such, it prevents R-loop formation by avoiding excessive RNAP IImediated transcription at DSB sites (Schwartz et al, 2012;Kwon et al, 2013;Hill et al, 2016). DDX5, a DEAD box-containing RNA helicase like DDX1, also unwinds RNA:DNA hybrids and R-loops at DSB sites (Xing et al, 2017;Mersaoui et al, 2019;Yu et al, 2020;Sessa et al, 2021). It binds to RNA that is transcribed in cis to a DSB to remove RNA-associated impediments and enable HR-directed repair (Yu et al, 2020;Sessa et al, 2021).…”

“…DDX5, a DEAD box-containing RNA helicase like DDX1, also unwinds RNA:DNA hybrids and R-loops at DSB sites (Xing et al, 2017;Mersaoui et al, 2019;Yu et al, 2020;Sessa et al, 2021). It binds to RNA that is transcribed in cis to a DSB to remove RNA-associated impediments and enable HR-directed repair (Yu et al, 2020;Sessa et al, 2021). Accordingly, loss of DDX5 results in the accumulation of polarized end deletions that specifically occur on those sides of DSBs that are actively transcribed and as such generate RNA species that can hinder the HR machinery (Yu et al, 2020).…”

“…Accordingly, loss of DDX5 results in the accumulation of polarized end deletions that specifically occur on those sides of DSBs that are actively transcribed and as such generate RNA species that can hinder the HR machinery (Yu et al, 2020). DDX5 interacts directly with the HR mediator BRCA2, which helps to enrich DDX5 at DSB sites, and which stimulates its RNA helicase activity in vitro (Sessa et al, 2021). Interestingly, like numerous other RBPs, DDX5 is then rapidly excluded again from damaged chromatin (Salton et al, 2010;Ha et al, 2011;Adamson et al, 2012;Polo et al, 2012;Britton et al, 2014;Altmeyer et al, 2015;Chen et al, 2018;Yu et al, 2020;Sessa et al, 2021).…”

“…DDX5 interacts directly with the HR mediator BRCA2, which helps to enrich DDX5 at DSB sites, and which stimulates its RNA helicase activity in vitro (Sessa et al, 2021). Interestingly, like numerous other RBPs, DDX5 is then rapidly excluded again from damaged chromatin (Salton et al, 2010;Ha et al, 2011;Adamson et al, 2012;Polo et al, 2012;Britton et al, 2014;Altmeyer et al, 2015;Chen et al, 2018;Yu et al, 2020;Sessa et al, 2021). This exclusion is ATM-and transcriptiondependent and requires the RNA-binding domain in DDX5 (Yu et al, 2020).…”