Abstract
Transcription-replication conflicts lead to DNA damage and genomic instability, which are closely related to human diseases. A major source of these conflicts is the formation of R-loops, which consist of an RNA-DNA hybrid and a displaced single-stranded DNA. Although these structures have been studied, many aspects of R-loop biology and R-loop-mediated genome instability remain unclear. Here, we demonstrate that thyroid hormone receptor-associated protein 3 (Thrap3) plays a critical role in regulating R-loop resolution. In cancer cells, Thrap3 interacts with DEAD-box helicase 5 (DDX5) and localizes to R-loops. Arginine-mediated methylation of DDX5 is required for its interaction with Thrap3, and the Thrap3-DDX5 axis induces the recruitment of 5’-3’ exoribonuclease 2 (XRN2) into R-loops. Loss of Thrap3 increases R-loop accumulation and DNA damage. These findings suggest that Thrap3 mediates resistance to cell death by preventing R-loop accumulation in cancer cells.
Highlights
R-loops are three-stranded nucleic acid structures consisting of a cotranscriptionally generated RNA-DNA hybrid and a displaced single-stranded DNA1–4
We found that Thrap[3] interacts with DEAD-box helicase 5 (DDX5) and that this interaction is followed by XRN2 recruitment for R-loop resolution
Using a proximity ligation assay with the S9.6 antibody, which can detect R-loops, and an anti-Thrap[3] antibody, we detected the colocalization of Thrap[3] with R-loops in the nuclear region; in addition, depletion of Thrap[3] using siRNA decreased the ligation signals in both cancer and normal (MEF) cells (Fig. 1a, Supplementary Fig. 1a, b)
Summary
R-loops are three-stranded nucleic acid structures consisting of a cotranscriptionally generated RNA-DNA hybrid and a displaced single-stranded DNA1–4. They have critical roles in a wide range of eukaryotic metabolic processes, such as chromosome segregation during mitosis, immunoglobulin class switching, DNA replication and repair, and transcription initiation/termination[3,4,5]. R-loops are involved in gene expression and rearrangement;[6] they threaten genome integrity, resulting in deleterious cellular effects. R-loops and subsequent collisions with replication forks is followed by DNA damage and genome instability[1,4,6,7]. Proper regulation of R-loops in cells is essential R-loops have been postulated to be causative factors for many human diseases, such as neurodegenerative diseases, cancer, and autoimmune diseases[2].
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