Dormant origins and fork protection mechanisms rescue sister forks arrested by transcription

Yeast Sen1Senataxin is a RNA/DNA helicase that preserves replication forks across RNA Polymerase II-transcribed genes by counteracting RNA:DNA hybrids accumulation. We show that in Sen1-depleted cells early forks clashing head-on with transcription halt, and impair progression of sister forks within the same replicon. Unsolved replication-transcription collisions trigger the local firing of dormant origins that rescue arrested forks. In sen1 mutants the MRX and Mrc1/Ctf4-complexes protect those forks clashing with transcription by preventing genotoxic fork-resection events mediated by the Exo1 nuclease. Hence, sister forks within the same replicon remain coupled when one of the two forks halts. This is different when forks encounter double strand breaks. Moreover, the local firing of dormant origins is not prevented by checkpoint activation but depends on delayed adjacent forks. Furthermore, a productive head-on clash between replication and transcription requires the tuning of origin firing and coordination between Sen1, the MRX and Mrc1/Ctf4-complexes and Exo1. Genome-Wide Analysis of DNA Synthesis by BrdU Immunoprecipitation on Tiling Microarrays (BrdU-IP-chip) was carried out as described (Viggiani et al., 2010), employing anti-BrdU antibody (MBL M1-11-3). Labelled probes were hybridized to Affymetrix S.cerevisiae Tiling 1.0 (P/N 900645) arrays and processed with TAS software.