R-Loop Physiology and Pathology: A Brief Review

Abstract

Physiological and pathological roles for R-loop structures continue to be discovered, and studies suggest that R-loops could contribute to human disease. R-loops are nucleic acid structures characterized by a DNA:RNA hybrid and displaced single-stranded DNA that occur in connection with transcription. R-loops form naturally and have been shown to be important for a number of physiological processes such as mitochondrial replication initiation, class switch recombination, DNA repair, modulating DNA topology, and regulation of gene expression. However, subsets of R-loops or persistent R-loops lead to DNA breaks, chromosome rearrangement, and genome instability. In addition, R-loops have been linked to human diseases, specifically neurological disorders and cancer. Of the large amount of research produced recently on R-loops, this review covers evidence for R-loop involvement in normal cellular physiology and pathophysiology, as well as describing factors that contribute to R-loop regulation.

Characterization of R-Loop–Interacting Proteins in Embryonic Stem Cells Reveals Roles in rRNA Processing and Gene Expression

“…In all, 49% of proteins (164 of 335) identified in our S9.6 co-IP/MS analysis are known RNA-binding proteins, including several known regulators of R-loop formation, suggesting a high proportion of hits are bona fide R-loop-interacting proteins. R-loops and other non-canonical DNA structures, such as G4s and cruciform DNA, have previously been shown to colocalize throughout the genome 74,75 . We J o u r n a l P r e -p r o o f therefore investigated if R-loop-interacting proteins identified in this study are known to bind to and regulate other DNA or RNA structures.…”

Pathways and signatures of mutagenesis at targeted DNA nicks

“…DNA2 possesses both 3' and 5' exonucleolytic activities, and interaction with BLM or WRN helicase can drive its 5' resection activity [8,[19][20][21], consistent with predominately 5' resection providing a source of asymmetry. The zone targeted for mutation coincides with that of the zone that forms an R-loop upon annealing to the 20 bp CRISPR gRNA, which may contribute to mutagenesis as R-loops are inherently unstable [49]. A single-stranded region approximately 20-30 nt in length is protected by a human RPA trimer [17,18], a dimension similar to that of the R-loop.…”

Proper control of R-loop homeostasis is required for maintenance of gene expression and neuronal function during aging

“…R-loops are three-stranded nucleic acid structures consisting of an RNA-DNA hybrid and a misplaced single-strand of DNA (Aguilera & García-Muse, 2012). They typically form during transcription in organisms ranging from yeast to humans (Aguilera & Gomez-Gonzalez, 2017) and play a significant role in normal cellular physiology, being required for the initiation of mitochondrial replication and class switch recombination (Mackay et al, 2020). Moreover, recent studies suggest that R-loops can dynamically regulate gene expression (Niehrs & Luke, 2020).…”

“…Loss of Top3β is associated with neurological disorders (Mackay et al, 2020) and has been shown to reduce lifespan in mice (Kwan & Wang, 2001).…”

“…Topoisomerase 3β (Top3β) is a member of the type IA superfamily of topoisomerases, which unwind negatively supercoiled DNA formed during transcription and replication, an activity that prevents R-loop formation (Drolet, 2006; Drolet et al, 1994, 1995). Loss of Top3β is associated with neurological disorders (Mackay et al, 2020) and has been shown to reduce lifespan in mice (Kwan & Wang, 2001).…”