Abstract
Aberrant transcription‐associated RNA:DNA hybrid (R‐loop) formation often causes catastrophic conflicts during replication, resulting in DNA double‐strand breaks and genomic instability. Preventing such conflicts requires hybrid dissolution by helicases and/or RNase H. Little is known about how such helicases are regulated. Herein, we identify DDX5, an RGG/RG motif‐containing DEAD‐box family RNA helicase, as crucial player in R‐loop resolution. In vitro, recombinant DDX5 resolves R‐loops in an ATP‐dependent manner, leading to R‐loop degradation by the XRN2 exoribonuclease. DDX5‐deficient cells accumulate R‐loops at loci with propensity to form such structures based on RNA:DNA immunoprecipitation (DRIP)‐qPCR, causing spontaneous DNA double‐strand breaks and hypersensitivity to replication stress. DDX5 associates with XRN2 and resolves R‐loops at transcriptional termination regions downstream of poly(A) sites, to facilitate RNA polymerase II release associated with transcriptional termination. Protein arginine methyltransferase 5 (PRMT5) binds and methylates DDX5 at its RGG/RG motif. This motif is required for DDX5 interaction with XRN2 and repression of cellular R‐loops, but not essential for DDX5 helicase enzymatic activity. PRMT5‐deficient cells accumulate R‐loops, resulting in increased formation of γH2AX foci. Our findings exemplify a mechanism by which an RNA helicase is modulated by arginine methylation to resolve R‐loops, and its potential role in regulating transcription.
Highlights
Aberrant transcription-associated RNA:DNA hybrid (R-loop) formation often causes catastrophic conflicts during replication, resulting in DNA double-strand breaks and genomic instability
We show that the methylation of the RNA helicase DDX5 by Protein arginine methyltransferase 5 (PRMT5) regulates its association with XRN2 to suppress R-loops at transcription termination regions and maintain genomic stability
The Cterminal RGG/RG motif of DDX5 was methylated by PRMT5 and required for R-loop resolution, as amino acid substitution of its arginines in the RGG/RG motif with lysines (RK) failed to reverse the accumulation of R-loops in siDDX5 cells
Summary
Aberrant transcription-associated RNA:DNA hybrid (R-loop) formation often causes catastrophic conflicts during replication, resulting in DNA double-strand breaks and genomic instability. Preventing such conflicts requires hybrid dissolution by helicases and/or RNase H. DDX5-deficient cells accumulate R-loops at loci with propensity to form such structures based on RNA:DNA immunoprecipitation (DRIP)-qPCR, causing spontaneous DNA double-strand breaks and hypersensitivity to replication stress. Protein arginine methyltransferase 5 (PRMT5) binds and methylates DDX5 at its RGG/RG motif This motif is required for DDX5 interaction with XRN2 and repression of cellular R-loops, but not essential for DDX5 helicase enzymatic activity. Our findings exemplify a mechanism by which an RNA helicase is modulated by arginine methylation to resolve R-loops, and its potential role in regulating transcription
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have