Single-molecule studies on co-transcriptional R-loop formation at DNA damage site.

Abstract

R-loop, a non-canonical nucleic acid structure, is composed of an RNA-DNA hybrid and a single-stranded DNA(ssDNA). This abnormal structure can form through transcription. R-loop can play a role in some biological processes such as gene regulation, but R-loop has been considered a putative risk factor causing genome instability. Recent studies, however, suggest that R-loop formation is also required for certain DNA double-strand break (DSB) repair processes implying even positive aspects of R-loop in light of the maintenance of genome integrity. Here, we show that E. coli RNA polymerases have an unexpected ability to create R-loop when they encounter DSB during transcription. The efficiency of DSB induced R-loop formation under single-round transcription was quantitatively measured. We also observed the propagation of RNA-DNA hybrids from break sites to upstream. These extended RNA-DNA hybrids hindered the next round of transcription. Furthermore, we studied whether other types of genetic lesions and abnormal structures could induce co-transcriptional R-loop formation.