Phosphorylation of Ku dictates DNA double-strand break (DSB) repair pathway choice in S phase

Kyung Jong Lee, Janapriya Saha, Jingxin Sun, Kazi R. Fattah, Shu Chi Wang, Burkhard Jakob, Linfeng Chi, Shih Ya Wang, Gisela Taucher-Scholz, Anthony J. Davis, David J. Chen

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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.

Original languageEnglish (US)
Pages (from-to)1732-1745
Number of pages14
JournalNucleic Acids Research
Volume44
Issue number4
DOIs
StatePublished - Dec 11 2015

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Double-Stranded DNA Breaks
Homologous Recombination
S Phase
Phosphorylation
DNA
Cell Cycle
1,2-di-(4-sulfamidophenyl)-4-butylpyrazolidine-3,5-dione

ASJC Scopus subject areas

  • Genetics

Cite this

Phosphorylation of Ku dictates DNA double-strand break (DSB) repair pathway choice in S phase. / Lee, Kyung Jong; Saha, Janapriya; Sun, Jingxin; Fattah, Kazi R.; Wang, Shu Chi; Jakob, Burkhard; Chi, Linfeng; Wang, Shih Ya; Taucher-Scholz, Gisela; Davis, Anthony J.; Chen, David J.

In: Nucleic Acids Research, Vol. 44, No. 4, 11.12.2015, p. 1732-1745.

Research output: Contribution to journalArticle

Lee, Kyung Jong ; Saha, Janapriya ; Sun, Jingxin ; Fattah, Kazi R. ; Wang, Shu Chi ; Jakob, Burkhard ; Chi, Linfeng ; Wang, Shih Ya ; Taucher-Scholz, Gisela ; Davis, Anthony J. ; Chen, David J. / Phosphorylation of Ku dictates DNA double-strand break (DSB) repair pathway choice in S phase. In: Nucleic Acids Research. 2015 ; Vol. 44, No. 4. pp. 1732-1745.
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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.",
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