Checkpoint-mediated DNA polymerase ε exonuclease activity curbing counteracts resection-driven fork collapse
2021
Summary DNA polymerase e (Pole) carries out high-fidelity leading strand synthesis owing to its exonuclease activity. Pole polymerase and exonuclease activities are balanced, because of partitioning of nascent DNA strands between catalytic sites, so that net resection occurs when synthesis is impaired. In vivo, DNA synthesis stalling activates replication checkpoint kinases, which act to preserve the functional integrity of replication forks. We show that stalled Pole drives nascent strand resection causing fork functional collapse, averted via checkpoint-dependent phosphorylation. Pole catalytic subunit Pol2 is phosphorylated on serine 430, influencing partitioning between polymerase and exonuclease active sites. A phosphormimetic S430D change reduces exonucleolysis in vitro and counteracts fork collapse. Conversely, non-phosphorylatable pol2-S430A expression causes resection-driven stressed fork defects. Our findings reveal that checkpoint kinases switch Pole to an exonuclease-safe mode preventing nascent strand resection and stabilizing stalled replication forks. Elective partitioning suppression has implications for the diverse Pole roles in genome integrity maintenance.
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