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Mol Cell. 2015 Apr 2;58(1):172-85. doi: 10.1016/j.molcel.2015.02.024. Epub 2015 Mar 26.

Differential phosphorylation of DNA-PKcs regulates the interplay between end-processing and end-ligation during nonhomologous end-joining.

Author information

1
Institute for Cancer Genetics, Department of Pathology and Cell Biology, College of Physicians and Surgeons, Columbia University, New York, NY 10032, USA.
2
Institute for Cancer Genetics, Department of Pathology and Cell Biology, College of Physicians and Surgeons, Columbia University, New York, NY 10032, USA; Pathobiology Graduate Program, Department of Pathology and Cell Biology, Columbia University, New York, NY 10032, USA.
3
University of Pittsburg, Hillman Cancer Center Research Pavilion Suite 2.6, 5117 Centre Avenue, Pittsburgh, PA 15213-1863, USA.
4
Division of Molecular Medicine and Genetics, Department of Internal Medicine, University of Michigan, Ann Arbor, MI 48109, USA.
5
Institute for Cancer Genetics, Department of Pathology and Cell Biology, College of Physicians and Surgeons, Columbia University, New York, NY 10032, USA; Division of Pediatric Oncology, Hematology and Stem Cell Transplantation, Department of Pediatrics, College of Physicians and Surgeons, Columbia University, New York, NY 10032, USA. Electronic address: sz2296@columbia.edu.

Abstract

Nonhomologous end-joining (NHEJ) is a major DNA double-strand break repair pathway that is conserved in eukaryotes. In vertebrates, NHEJ further acquires end-processing capacities (e.g., hairpin opening) in addition to direct end-ligation. The catalytic subunit of DNA-PK (DNA-PKcs) is a vertebrate-specific NHEJ factor that can be autophosphorylated or transphosphorylated by ATM kinase. Using a mouse model expressing a kinase-dead (KD) DNA-PKcs protein, we show that ATM-mediated transphosphorylation of DNA-PKcs regulates end-processing at the level of Artemis recruitment, while strict autophosphorylation of DNA-PKcs is necessary to relieve the physical blockage on end-ligation imposed by the DNA-PKcs protein itself. Accordingly, DNA-PKcs(KD/KD) mice and cells show severe end-ligation defects and p53- and Ku-dependent embryonic lethality, but open hairpin-sealed ends normally in the presence of ATM kinase activity. Together, our findings identify DNA-PKcs as the molecular switch that coordinates end-processing and end-ligation at the DNA ends through differential phosphorylations.

PMID:
25818648
PMCID:
PMC4415111
DOI:
10.1016/j.molcel.2015.02.024
[Indexed for MEDLINE]
Free PMC Article

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