Abstract
R-loop, a three-stranded nucleic acid structure consisting of the RNA:DNA hybrid and the displaced singlestranded DNA, is crucial for many cellular processes but could be a threat to genome integrity if dysregulated. The homeostasis of R-loops is governed by various factors including helicases, nucleases, and chromatin remodelers. Since there are many excellent reviews about R-loops, we focus on discussing how R-loop homeostasis is regulated via nucleic acid and protein modifications. We summarize how RNA modifications such as N6-methyladenosine (m6A), N5-methylcytosine (m5C), and potentially 3-methylcytidine (m3C), alongside DNA modifications such as deamination, methylation, and oxidation, influence R-loop dynamics. Moreover, we discuss how protein modifications, including ubiquitination, SUMOylation, acetylation, methylation, and phosphorylation, modulate the activity, stability, or recruitment of R-loop processing factors. Importantly, these modifications often interact with each other and exhibit context-dependent roles, either promoting R-loop formation or facilitating resolution. Elucidating how these chemical codes orchestrate R-loop homeostasis will facilitate our understanding of the mechanisms governing R-loop homeostasis and could provide some insights into genome maintenance, gene expression, and pathogenesis caused by R-loop dysregulation.