title: Phosphorylation of Ku dictates DNA double-strand break (DSB) repair pathway choice in S phase.
abstract: Multiple DNA double-strand break (DSB) repair pathways are active in S phase of the cell cycle; however, DSBs are primarily repaired by homologous recombination (HR) in this cell cycle phase. As the non-homologous end-joining (NHEJ) factor, Ku70/80 (Ku), is quickly recruited to DSBs in S phase, we hypothesized that an orchestrated mechanism modulates pathway choice between HR and NHEJ via displacement of the Ku heterodimer from DSBs to allow HR. Here, we provide evidence that phosphorylation at a cluster of sites in the junction of the pillar and bridge regions of Ku70 mediates the dissociation of Ku from DSBs. Mimicking phosphorylation at these sites reduces Ku's affinity for DSB ends, suggesting that phosphorylation of Ku70 induces a conformational change responsible for the dissociation of the Ku heterodimer from DNA ends. Ablating phosphorylation of Ku70 leads to the sustained retention of Ku at DSBs, resulting in a significant decrease in DNA end resection and HR, specifically in S phase. This decrease in HR is specific as these phosphorylation sites are not required for NHEJ. Our results demonstrate that the phosphorylation-mediated dissociation of Ku70/80 from DSBs frees DNA ends, allowing the initiation of HR in S phase and providing a mechanism of DSB repair pathway choice in mammalian cells.
authors.affiliation: Division of Molecular Radiation Biology, Department of Radiation Oncology, University of Texas Southwestern Medical Center, 2201 Inwood Rd, Dallas, Texas 75390, USA.
authors.forename: Kyung-Jong
authors.initials: KJ
authors.lastname: Lee
authors.affiliation: Division of Molecular Radiation Biology, Department of Radiation Oncology, University of Texas Southwestern Medical Center, 2201 Inwood Rd, Dallas, Texas 75390, USA.
authors.forename: Janapriya
authors.initials: J
authors.lastname: Saha
authors.affiliation: Division of Molecular Radiation Biology, Department of Radiation Oncology, University of Texas Southwestern Medical Center, 2201 Inwood Rd, Dallas, Texas 75390, USA.
authors.forename: Jingxin
authors.initials: J
authors.lastname: Sun
authors.affiliation: Division of Molecular Radiation Biology, Department of Radiation Oncology, University of Texas Southwestern Medical Center, 2201 Inwood Rd, Dallas, Texas 75390, USA.
authors.forename: Kazi R
authors.initials: KR
authors.lastname: Fattah
authors.affiliation: Division of Molecular Radiation Biology, Department of Radiation Oncology, University of Texas Southwestern Medical Center, 2201 Inwood Rd, Dallas, Texas 75390, USA.
authors.forename: Shu-Chi
authors.initials: SC
authors.lastname: Wang
authors.affiliation: Department of Biophysics; GSI Helmholtzzentrum für Schwerionenforschung GmbH, Planckstraße 1, Darmstadt, Germany.
authors.forename: Burkhard
authors.initials: B
authors.lastname: Jakob
authors.affiliation: Division of Molecular Radiation Biology, Department of Radiation Oncology, University of Texas Southwestern Medical Center, 2201 Inwood Rd, Dallas, Texas 75390, USA.
authors.forename: Linfeng
authors.initials: L
authors.lastname: Chi
authors.affiliation: Division of Molecular Radiation Biology, Department of Radiation Oncology, University of Texas Southwestern Medical Center, 2201 Inwood Rd, Dallas, Texas 75390, USA.
authors.forename: Shih-Ya
authors.initials: SY
authors.lastname: Wang
authors.affiliation: Department of Biophysics; GSI Helmholtzzentrum für Schwerionenforschung GmbH, Planckstraße 1, Darmstadt, Germany.
authors.forename: Gisela
authors.initials: G
authors.lastname: Taucher-Scholz
authors.affiliation: Division of Molecular Radiation Biology, Department of Radiation Oncology, University of Texas Southwestern Medical Center, 2201 Inwood Rd, Dallas, Texas 75390, USA.
authors.forename: Anthony J
authors.initials: AJ
authors.lastname: Davis
authors.affiliation: Division of Molecular Radiation Biology, Department of Radiation Oncology, University of Texas Southwestern Medical Center, 2201 Inwood Rd, Dallas, Texas 75390, USA david.chen@utsouthwestern.edu.
authors.forename: David J
authors.initials: DJ
authors.lastname: Chen
mesh_terms: D000818:Animals; D034961:Antigens, Nuclear / Q000235:genetics*; D053903:DNA Breaks, Double-Stranded; D004249:DNA Damage / Q000235:genetics; D059766:DNA End-Joining Repair / Q000235:genetics*; D004260:DNA Repair / Q000235:genetics; D004268:DNA-Binding Proteins / Q000235:genetics*; D005347:Fibroblasts / Q000378:metabolism; D045325:HCT116 Cells; D059765:Homologous Recombination; D006801:Humans; D000072200:Ku Autoantigen; D051379:Mice; D016196:S Phase / Q000235:genetics*; D015398:Signal Transduction
chemical_list: D034961:Antigens, Nuclear; D004268:DNA-Binding Proteins; C000606509:Xrcc6 protein, human; C000606510:Xrcc6 protein, mouse; D000072200:Ku Autoantigen