Genome-wide distribution of RNA-DNA hybrids identifies RNase H targets in tRNA genes, retrotransposons and mitochondria

During transcription the nascent RNA can invade the DNA template, forming extended RNA-DNA duplexes (R-loops). Here we employ ChIP-seq in strains expressing or lacking RNase H to map targets of RNase H activity throughout budding yeast genome. In wild-type strains, R-loops were readily detected over the 35S rDNA region transcribed by Pol I and over the 5S rDNA transcribed by Pol III. In strains lacking RNase H activity, R-loops were elevated over other Pol III genes notably tRNAs, SCR1 and U6 snRNA, and were also associated with the cDNAs of endogenous TY1 retrotransposons, which showed increased rates of mobility to the 5?-flanking regions of tRNA genes. Unexpectedly, R-loops were also associated with mitochondrial genes in the absence of RNase H1, but not of RNase H2. Finally, R-loops were detected on highly expressed protein-coding genes in the wild-type, notably over the second exon of spliced ribosomal protein genes. Overall design: ChIP-seq of RNA-DNA hybrids using antibody S9.6