Sen1 Is Recruited to Replication Forks via Ctf4 and Mrc1 and Promotes Genome Stability

DNA replication and RNA transcription compete for the same substrate during S phase. Cells have evolved several mechanisms to minimize such conflicts. Here, we identify the mechanism by which the transcription termination helicase Sen1 associates with replisomes. We show that the N terminus of Sen1 is both sufficient and necessary for replisome association and that it binds to the replisome via the components Ctf4 and Mrc1. We generated a separation of function mutant, sen1-3, which abolishes replisome binding without affecting transcription termination. We observe that the sen1-3 mutants show increased genome instability and recombination levels. Moreover, sen1-3 is synthetically defective with mutations in genes involved in RNA metabolism and the S phase checkpoint. RNH1 overexpression suppresses defects in the former, but not the latter. These findings illustrate how Sen1 plays a key function at replication forks during DNA replication to promote fork progression and chromosome stability. [Display omitted] •The N-terminal domain of Sen1 mediates replisome association•Ctf4 and Mrc1 bind to Sen1 and promote its recruitment to the replisome•The sen1-3 allele abrogates replisome binding, but not transcription termination•sen1-3 cells are sensitive to high levels of R-loops and defects of S phase checkpoint Appanah et al. identify the transcription termination helicase Sen1 as a bona fide component of the replisome. Sen1 binds the replisome via its N-terminal domain and Ctf4 and Mrc1. The allele sen1-3 breaks this interaction without affecting transcription termination. sen1-3 cells show sensitivity to R-loops levels and increased genomic instability.