Histone deposition promotes recombination-dependent replication at arrested forks

Replication stress poses a serious threat to genome stability. Recombination-Dependent-Replication (RDR) promotes DNA synthesis resumption from arrested forks. Despite the identification of chromatin restoration pathways after DNA repair, crosstalk coupling RDR and chromatin assembly is largely unexplored. The fission yeast Chromatin Assembly Factor-1, CAF-1, is known to promote RDR. Here, we addressed the contribution of histone deposition to RDR. We expressed a mutated histone, H3-H113D, to genetically alter replication-dependent chromatin assembly by destabilizing (H3-H4)(2) tetramer. We established that DNA synthesis-dependent histone deposition, by CAF-1 and Asf1, promotes RDR by preventing Rqh1-mediated disassembly of joint-molecules. The recombination factor Rad52 promotes CAF-1 binding to sites of recombination-dependent DNA synthesis, indicating that histone deposition occurs downstream Rad52. Histone deposition and Rqh1 activity act synergistically to promote cell resistance to camptothecin, a topoisomerase I inhibitor that induces replication stress. Moreover, histone deposition favors non conservative recombination events occurring spontaneously in the absence of Rqh1, indicating that the stabilization of joint-molecules by histone deposition also occurs independently of Rqh1 activity. These results indicate that histone deposition plays an active role in promoting RDR, a benefit counterbalanced by stabilizing at-risk joint-molecules for genome stability. Author summary DNA replication occurs in the context of DNA packaged into chromatin. At replication fork, the parental chromatin is evicted and re-assembled, together with newly synthetized histones, onto newly replicated DNA thanks to a network of histone chaperones. The progression of replication forks is often interrupted by a plethora of fork obstacles that affect replisome functionality. Such dysfunctional forks are fragile structures prone to chromosomal rearrangements and chromatin changes, leading to human diseases including cancer. Homologous recombination plays a crucial role in the recovery of dysfunctional forks. We have genetically altered the process of histone deposition-coupled to DNA synthesis to interrogate the contribution of chromatin assembly during fork recovery by homologous recombination. We found that histone deposition acts during fork recovery in a manner that recombination structures are protected from disassembly by helicases such as Rqh1. Histone deposition-coupled to fork recovery favors deletion-type recombinant. Chromatin assembly during fork recovery requires only a subset of the histone chaperones normally acting at canonical forks, namely CAF-1 and Asf1. CAF-1 associates to sites of recombination-dependent DNA synthesis during fork recovery. We propose that restarted forks remain coupled to histone deposition, a benefit counterbalanced by the risk to stabilize recombination intermediates detrimental to genome stability.