Abstract: Co-transcriptional R-loops are formed in yeast mutants of the THO complex, which functions at the interface between transcription and mRNA export. Despite the relevance of R-loops in transcription-associated recombination, the mechanisms by which they trigger recombination are still elusive. In order to understand how R-loops compromise genome stability we have analyzed the genetic interaction of THO with 26 genes involved in replication, S-phase checkpoint, DNA repair and chromatin remodeling. We found a synthetic growth defect in double null mutants of THO and S-phase checkpoint factors, such as the RFC- and PCNA-like complexes. Under replicative stress, R-loop forming THO null mutants require functional S-phase checkpoint functions but not double-strand break (DSB) repair functions for survival. Furthermore, R-loop forming hpr1Delta mutants display replication fork-progression impairment at actively transcribed chromosomal regions and trigger Rad53 phosphorylation. We conclude that R-loop-mediated DNA damage activates the S-phase checkpoint, which is required for cell survival of THO mutants under replicative stress. In light of these results, we propose a model in which R-loop-mediated recombination is explained by template switching.