Mapping Native R-Loops Genome-wide Using a Targeted Nuclease Approach

Yan, Qingqing; Shields, Emily; Bonasio, Roberto; Sarma, Kavitha

doi:10.1016/j.celrep.2019.09.052

Cited by 77 publications

(79 citation statements)

References 50 publications

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“…In addition, only 5.5%–15.8% of R-loops were detected in the intergenic region (Fig. 1 B), which implies that the majority of R-loops preferentially form surrounding gene regions, which is consistent with previous reports [ 3 , 24 , 51 ]. In addition, R-loops formed most frequently in promoter regions across species (Fig.…”

Section: Resultssupporting

confidence: 92%

Association analysis of repetitive elements and R-loop formation across species

2021

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Background Although recent studies have revealed the genome-wide distribution of R-loops, our understanding of R-loop formation is still limited. Genomes are known to have a large number of repetitive elements. Emerging evidence suggests that these sequences may play an important regulatory role. However, few studies have investigated the effect of repetitive elements on R-loop formation. Results We found different repetitive elements related to R-loop formation in various species. By controlling length and genomic distributions, we observed that satellite, long interspersed nuclear elements (LINEs), and DNA transposons were each specifically enriched for R-loops in humans, fruit flies, and Arabidopsis thaliana, respectively. R-loops also tended to arise in regions of low-complexity or simple repeats across species. We also found that the repetitive elements associated with R-loop formation differ according to developmental stage. For instance, LINEs and long terminal repeat retrotransposons (LTRs) are more likely to contain R-loops in embryos (fruit fly) and then turn out to be low-complexity and simple repeats in post-developmental S2 cells. Conclusions Our results indicate that repetitive elements may have species-specific or development-specific regulatory effects on R-loop formation. This work advances our understanding of repetitive elements and R-loop biology.

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Section: Resultssupporting

confidence: 92%

Association analysis of repetitive elements and R-loop formation across species

2021

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“…Lastly, we examined whether TonEBP recruitment and m6A methylation would be reduced when R-loop formation was blocked using actinomycin D ( 28 , 35 , 36 ). To avoid unexpected change of protein expression level, cells were treated with high concentration of actinomycin D (10 μM) for a short time (10 min).…”

Section: Resultsmentioning

confidence: 99%

TonEBP recognizes R-loops and initiates m6A RNA methylation for R-loop resolution

Kang

Cheon

Park

et al. 2020

Nucleic Acids Research

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R-loops are three-stranded, RNA–DNA hybrid, nucleic acid structures produced due to inappropriate processing of newly transcribed RNA or transcription-replication collision (TRC). Although R-loops are important for many cellular processes, their accumulation causes genomic instability and malignant diseases, so these structures are tightly regulated. It was recently reported that R-loop accumulation is resolved by methyltransferase-like 3 (METTL3)-mediated m6A RNA methylation under physiological conditions. However, it remains unclear how R-loops in the genome are recognized and induce resolution signals. Here, we demonstrate that tonicity-responsive enhancer binding protein (TonEBP) recognizes R-loops generated by DNA damaging agents such as ultraviolet (UV) or camptothecin (CPT). Single-molecule imaging and biochemical assays reveal that TonEBP preferentially binds a R-loop via both 3D collision and 1D diffusion along DNA in vitro. In addition, we find that TonEBP recruits METTL3 to R-loops through the Rel homology domain (RHD) for m6A RNA methylation. We also show that TonEBP recruits RNaseH1 to R-loops through a METTL3 interaction. Consistent with this, TonEBP or METTL3 depletion increases R-loops and reduces cell survival in the presence of UV or CPT. Collectively, our results reveal an R-loop resolution pathway by TonEBP and m6A RNA methylation by METTL3 and provide new insights into R-loop resolution processes.

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“…The parallel is possible. With S9.6, RNase H and more recently MapR 22 established that the GC content is important in the recognition of the R-loop by antibodies and in particular by RNaseH 23 . Alternatively, here unlike the previous methods, R-loop detection without protein of DNA/RNA hybrids in normal cells (sperm and testes) reveals, the stable structure of DNA-RNA hybrids which are not particularly rich in CpG.…”

Section: Discussionmentioning

confidence: 99%

DNA-RNA hybrid (R-loop): from a unified picture of the mammalian telomere to the genome-wide

Rassoulzadegan

Sharifi‐Zarchi

Kianmehr

2020

Preprint

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SummaryLocal three-stranded DNA-RNA hybrid regions of the genomes (R-loops) have been detected either by binding of a monoclonal antibody (DRIP assay) or by enzymatic recognition by RNaseH. Such a structure has been postulated for the mouse and human telomeres, clearly suggested by the identification of the complementary RNA TERRA. It was, however, evidenced by DRIP exclusively in human cancer and not in normal human nor in any mouse cell. Based on the observation that in several fractionation procedures, DNA-RNA hybrids copurify with double-stranded DNA, we developed a preparative approach that allows detection of stable DNA-RNA triplexes in a complex genome, their physical isolation and their RNA nucleotide sequence. We then define in the normal mouse and human genomes the notion of terminal R-loop complexes including TERRA molecules synthesized from local promoters of every chromosome. In addition to the telomeric structures since, the finding of the RNA peak, applied in addition to the whole murine sperm genomes, has highlighted a reproducible profile of the R-loop complexes from entire genome suggesting a defined profile for a future generation.

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Mapping Native R-Loops Genome-wide Using a Targeted Nuclease Approach

Cited by 77 publications

References 50 publications

Association analysis of repetitive elements and R-loop formation across species

Association analysis of repetitive elements and R-loop formation across species

TonEBP recognizes R-loops and initiates m6A RNA methylation for R-loop resolution

DNA-RNA hybrid (R-loop): from a unified picture of the mammalian telomere to the genome-wide

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