Journal of Biological Chemistry
Volume 280, Issue 15, 15 April 2005, Pages 14709-14715
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DNA: Replication, Repair, and Recombination
Cell Cycle Dependence of DNA-dependent Protein Kinase Phosphorylation in Response to DNA Double Strand Breaks*

https://doi.org/10.1074/jbc.M408827200Get rights and content
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DNA-dependent protein kinase (DNA-PK), consisting of Ku and DNA-PKcs subunits, is the key component of the non-homologous end-joining (NHEJ) pathway of DNA double strand break (DSB) repair. Although the kinase activity of DNA-PKcs is essential for NHEJ, thus far, no in vivo substrate has been conclusively identified except for an autophosphorylation site on DNA-PKcs itself (threonine 2609). Here we report the ionizing radiation (IR)-induced autophosphorylation of DNA-PKcs at a novel site, serine 2056, the phosphorylation of which is required for the repair of DSBs by NHEJ. Interestingly, IR-induced DNA-PKcs autophosphorylation is regulated in a cell cycle-dependent manner with attenuated phosphorylation in the S phase. In contrast, DNA replication-associated DSBs resulted in DNA-PKcs autophosphorylation and localization to DNA damage sites. These results indicate that although IR-induced DNA-PKcs phosphorylation is attenuated in the S phase, DNA-PKcs is preferentially activated by the physiologically relevant DNA replication-associated DSBs at the sites of DNA synthesis.

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*

This work was supported by the United States Department of Energy under contract DE-AC03-76SF00098 and by National Institutes of Health Grants CA50519, CA86936, and PO1-CA92584. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

Supported by postdoctoral fellowships from the Canadian Institutes of Health Research (CIHR) and the Alberta Heritage Foundation for Medical Research.

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Recipient of Concept Award DAMD17-03-1-0635 from the Department of Defense Breast Cancer Research Program.

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Recipient of Career Development Award DAMD17-00-1-0146 from the Department of Defense Breast Cancer Research Program.