ATR Signaling Uncouples the Role of RAD51 Paralogs in Homologous Recombination and Replication Stress Response

ATR kinase-mediated replication checkpoint is vital for genome maintenance following replication stress. Previously, we showed that XRCC2-RAD51D (DX2) sub-complex of RAD51 paralogs restrains active DNA synthesis during dNTP alterations, in a manner dependent on ATR-mediated phosphorylation of XRCC2. Here, we find that unrestrained fork progression in XRCC2 deficiency and phosphorylation defect causes replication-associated errors, subsequently resulting in genome-wide double-strand breaks (DSBs) and early activation of ATM signaling. Cells defective in XRCC2 phosphorylation exhibit ATM/ATR-mediated early activation of XRCC3 during perturbed replication, which facilitates recombination-mediated repair of the post-replicative DNA damage and thereby promotes cell viability. Collectively, our findings identify collaborative roles of RAD51 paralog complexes during replication stress and reveal their differential regulation by ATR signaling to promote cell survival and genome integrity.