Fig. 3

Genome-wide detection of R-loops during meiosis by ssDRIP-seq. (A) The number of overlapping ssDRIP-seq peaks from two biological repeats during yeast meiosis (0 h, 0.5 h, and 2 h in traditional synchronized SK1 strains [41]; 6 h, pachytene stage, Pac; 7.5 h, metaphase I, MI; and 8 h, anaphase I, AI in NDT80-induced synchronized SK1 strains [39]). The ssDRIP-seq peaks were analysed by peak calling strategies of MACS2 with default settings to call narrow R-loop peaks [63]. (B) Snapshot of the ssDRIP-seq data during yeast sporulation (0 h, 0.5 h, 2 h, Pac, MI, AI), including R-loop (grey), wR-loop (blue) and cR-loop (red) in a representative genomic region (chrI:70,000–150,000). (C) R-loop loads onto meiotic chromosomes during prophase I. Yeast cells were incubated in SPM and harvested at 6 h, and meiotic chromosomes were spread for immunofluorescence (Zip1-GFP, green; S9.6, red). Nuclei were stained with DAPI (blue). RNase H treatment served as a negative control. (D) The genomic distribution of R-loops during meiosis (0 h, 0.5 h, 2 h, Pac, MI, AI) that were identified by ChIPseeker. Various genomic regions are colour coded according to the labels at the bottom (blue, promoter; green, 1st exon; yellow, other exon; black, intron; orange, downstream [≤ 300 bp]; red, distal intergenic [> 500 bp from TSS and > 300 bp from TES]). (E) DNA motif in the peak regions of unstranded R-loops, wR-loops and cR-loops during meiosis (0 h, 0.5 h, 2 h, Pac, MI, AI) that were identified by MEME-ChIP. E-values are provided on the right