ATM safeguards DNA replication by restraining pathological repriming at endogenous base lesions.

Abstract

Poly(ADP-ribose) polymerase inhibitors (PARPi) exploit homologous recombination (HR) deficiency in BRCA1/2-mutant cancers to induce synthetic lethality. PARPi also kill ataxia telangiectasia-mutated (ATM)-deficient cells, but the underlying mechanism has remained unclear. Here, we define the mechanism of PARPi cytotoxicity in mammalian ATM-deficient cells, uncovering a critical role for ATM in DNA replication. In the absence of ATM, unrestrained PRIMPOL-dependent repriming at oxidative base lesions generates post-replicative DNA gaps that activate PARP. This defect is driven by aberrant BRCA1-A recruitment to stalled replication forks in ATM-deficient cells, suppressing HLTF/ZRANB3-mediated replication fork slowing. These DNA gaps require HR for repair and underlie PARPi synthetic lethality. Suppressing repriming or base excision repair, or reducing oxidative stress, alleviates these defects. Our findings reveal how spontaneous oxidative DNA damage synergizes with dysregulated replication to drive PARPi sensitivity, establishing a paradigm of post-replicative repair addiction in ATM-deficient cells and linking oxidative DNA damage to genome instability in ataxia telangiectasia.