Transcription-Replication Conflict Resolution by Nuclear RNA Interference [sRNA-seq]

RNA interference (RNAi) plays an important role in the nucleus critical to genome maintenance and heterochromatin silencing. In the fission yeast Schizosaccharomyces pombe, Dicer (Dcr1) also plays a critical role in releasing RNA polymerase II (Pol II) to limit transcription-replication (T-R) stress, but a mechanistic insight is still lacking. Here, we show that Dcr1 resolves T-R conflicts by processing promoter-proximal co-transcriptional R-loops, facilitating replication progression and limiting DNA damage. We found in RNase H-deficient cells, which accumulated pathological R-loops, that further mutating Dcr1 caused hypersensitivity to genotoxic stress. Genetic evidence implicates Dcr1 in regulating nascent transcription, with the helicase domain providing an additional catalytic function in genome stability reminiscent of archaeal helicase homologs involved in replication. In the absence of Dcr1, DNA breaks accumulate around transcription start sites (TSSs), termination sites (TTSs) and pause sites, consistent with replication collision, and DNA replication processivity and speed are impaired in a manner dependent on transcriptional activity and directionality. Argonaute (Ago1) is implicated in dcr1?-induced genome instability, as its deletion suppressed the genotoxic stress phenotype of dcr1?. Our results demonstrate a novel nuclear function of Dcr1, likely ancestral to the RNAi pathway. Overall design: small RNA sequencing (sRNA-seq) of WT, dcr1?, rnh1? rnh201?, and the triple mutant rnh1? rnh201? dcr1?