Genomic distribution of R-loop peaks during spermatogenesis. A: Pie charts showing the genomic distribution variation of R-loop peaks during spermatogenesis. SPG, spermatogonia; L/Z, leptotene/zygotene; P/D, pachytene/diplotene; MII, secondary spermatocytes. B: Genomic metaplots showing relative signal intensity comparison of R-loops across the 2 kb window around gene bodies in spermatogonia and spermatocytes. R-loop signals of different stages were normalized based on library sizes and spike-in scaling factors. C: Alluvial plots showing the global dynamics of genes with R-loop and genes without R-loops during spermatocyte development. Each line represents an R-loop gene, defined as a gene-detected R-loop peak in at least one analyzed stage. D: Box plots showing the expression level comparison of different gene clusters grouped by R-loop variation between adjacent stages in meiotic progress. E: Scatter plots showing the comparison of signal intensities of R-loop peaks between leptotene/zygotene and pachytene/diplotene spermatocytes. F: Volcano plots showing differentially expressed genes between leptotene/zygotene and pachytene spermatocytes. Genes upregulated (FC > 2; FDR < 0.05) and downregulated (FC < 1/2; FDR < 0.05) in pachytene spermatocytes are colored red and blue, respectively. G: Venn diagrams showing the overlap of R-loop accumulated genes and transcriptionally upregulated R-loop genes in leptotene/zygotene to pachytene stage. H: Venn diagrams showing the overlap of R-loop decreased genes and transcriptionally downregulated R-loop genes in leptotene/zygotene to pachytene stage. I: Venn diagrams showing the overlap of H3K4me3 peaks in genes promoter regions and R-loop peaks in leptotene/zygotene spermatocytes (Fisher’s exact test; ***P < 0.001). J: Venn diagrams showing the overlap of H3K4me3 peaks in genes promoter regions and R-loop peaks in pachytene spermatocytes (Fisher’s exact test; ***P < 0.001). K: Example snapshot of the relative distribution of the R-loop, H3K4me3, and downstream transcriptional level. (For interpretation of the references to color in this figure legend, the reader is referred to the web version of this article.)