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A series of critical pathways are responsible for the detection, signaling, and restart of replication forks that encounter blocks during S-phase progression. Small base lesions may obstruct replication fork progression and processing, but the link between repair of small lesions and replication forks is unclear. In this study, we investigated a hypothesized role for DNA-PK, an important enzyme in DNA repair, in cellular responses to DNA replication stress. The enzyme catalytic subunit DNA-PKcs was phosphorylated on S2056 at sites of stalled replication forks in response to short hydroxyurea treatment. Using DNA fiber experiments, we found that catalytically active DNA-PK was required for efficient replication restart of stalled forks. Furthermore, enzymatically active DNA-PK was also required for PARP-dependent recruitment of XRCC1 to stalled replication forks. This activity was enhanced by preventing Mre11-dependent DNA end resection, suggesting that XRCC1 must be recruited early to an unresected stalled fork. We also found that XRCC1 was required for effective restart of a subset of stalled replication forks. Overall, our work suggested that DNA-PK and PARP-dependent recruitment of XRCC1 is necessary to effectively protect, repair, and restart stalled replication forks, providing new insight into how genomic stability is preserved.

Original publication

DOI

10.1158/0008-5472.CAN-15-0608

Type

Journal article

Journal

Cancer Res

Publication Date

01/03/2016

Volume

76

Pages

1078 - 1088

Keywords

Casein Kinase II, Cell Line, DNA Repair, DNA Replication, DNA-Activated Protein Kinase, DNA-Binding Proteins, Humans, MRE11 Homologue Protein, Nuclear Proteins, Poly (ADP-Ribose) Polymerase-1, Poly(ADP-ribose) Polymerases, X-ray Repair Cross Complementing Protein 1