The R-loop is a common chromatin feature of the Arabidopsis genome

Abstract

R-loops are functional structures in chromatin comprising one single-stranded DNA and a DNA:RNA hybrid. Here, we report ssDRIP-seq, a single-strand DNA ligation-based library preparation technique for genome-wide identification of R-loops. When applied in Arabidopsis, ssDRIP-seq exhibits high efficiency, low bias and strand specificity. We found that Arabidopsis R-loops are enriched by both AT and GC skews, and are formed in the sense and antisense orientations. R-loops are strongly enriched in gene promoters and gene bodies, and are highly associated with noncoding RNA and repetitive genomic regions. Furthermore, R-loops are negatively correlated with CG DNA hypermethylation, and are prevalent in regions with multiple chromatin modifications, showing strong correlations with the activated and repressed gene loci. Our analyses indicate that R-loops are common features in the Arabidopsis genome and suggest that the R-loops play diverse roles in genome organization and gene regulation, thereby providing insights into plant nuclear genome formation and function.

The Affinity of the S9.6 Antibody for Double-Stranded RNAs Impacts the Accurate Mapping of R-Loops in Fission Yeast

“…2c and see [18]), were an exception to this low-expression rule for class B R-loops. The presence of R-loops at tDNAs confirms our previous observations [18] and observations made in budding yeast [7,20,21] and A. thaliana [9]. The low-level R-loop formation at tDNAs in wild-type cells is also consistent with the high recruitment of RNase H1 and Senataxin (Sen1) at these loci in fission yeast [18].…”

“…Altogether, our improved R-loop maps show that 5% of the fission yeast genome can exist under an R-loop form in unperturbed cells, an observation consistent with findings in mammals and plants [8,9]. Under these conditions, R-loop formation is principally a property of highly transcribed genes (class A).…”

“…This shows that one of the roles of endogenous RNase H enzymes is to suppress low-level R-loop formation over a significant portion of the genome. Our data, combined with observations from mammalian cells and A. thaliana [8,9,17,25], reinforce the idea that genuine R-loops share universal hallmarks, including strand specificity, co-directionality with transcription, co-dependence with transcription levels and a strong relationship with favorable DNA sequence features such as GC skew (Fig. 3).…”

“…We speculate that at least a fraction of these dsRNAs remain associated with chromatin as was suggested previously for heterochromatin transcripts [24], which might explain why they have been previously detected using DRIP. Interestingly, lack of a significant enrichment for R-loops at terminator regions was also recently observed in A. thaliana [9]. The situation in S. pombe and A. thaliana is therefore different from human and mouse where genuine R-loops form at a subset of terminator regions [8,17,25–28].…”

“…Recent maps have shown that R-loops can occupy between 5% and 10% of the genome in yeast, vertebrates and Arabidopsis thaliana [7–9]. R-loop formation has been associated with replication stress and genome instability, and it was recently proposed that defective control of R-loop formation could contribute to a number of human pathologies (reviewed in Refs.…”