Sources, resolution and physiological relevance of R-loops and RNA–DNA hybrids

RNA–DNA hybrids are generated during transcription, DNA replication and DNA repair and are crucial intermediates in these processes. When RNA–DNA hybrids are stably formed in double-stranded DNA, they displace one of the DNA strands and give rise to a three-stranded structure called an R-loop. R-loops are widespread in the genome and are enriched at active genes. R-loops have important roles in regulating gene expression and chromatin structure, but they also pose a threat to genomic stability, especially during DNA replication. To keep the genome stable, cells have evolved a slew of mechanisms to prevent aberrant R-loop accumulation. Although R-loops can cause DNA damage, they are also induced by DNA damage and act as key intermediates in DNA repair such as in transcription-coupled repair and RNA-templated DNA break repair. When the regulation of R-loops goes awry, pathological R-loops accumulate, which contributes to diseases such as neurodegeneration and cancer. In this Review, we discuss the current understanding of the sources of R-loops and RNA–DNA hybrids, mechanisms that suppress and resolve these structures, the impact of these structures on DNA repair and genome stability, and opportunities to therapeutically target pathological R-loops.