DNA double-strand breaks (DSBs) are critical lesions as they can generate genome rearrangements. This project has been focused on the roles of DNA dependent protein complex (DNA-PK) and its interactions with other DNA repair factors involved in early DSB responses. With our extensive and persistent effort during the past funding periods we begin to understand the molecular mechanisms whereby DNA-PK complex acts in DSB sensing, signaling and repair. Our results strongly support the hypothesis that DNA-PK is required for the initial phase of DSB sensing throughout the cell cycle upon DNA damage. Furthermore, the kinase activity of DNA-PKcs plays a pivotal role in modulating the cellular choice of DSB repair pathways between nonhomolgous end joining (NHEJ) and homologous recombination (HR). In this renewal application, we propose to test these hypotheses by employing cellular, molecular and transgenic mouse model approaches. Especially, we will utilize newly established in vivo imaging facility to dissect the temporal relationship among DNA-PK and other DSB sensing complex at DSB sites in live cells. Our research not only will provide a fundamental understanding of the mechanism of DSB repair, but also will unveil mechanism-based targets for the development of potential radiosensitization tools for cancer therapy.
The specific aims of the renewal application are: 1) To dissect the mechanism by which DNA-PKcs is activated at sites of DNA DSBs and the role of DNA-PK complex (DNA-PKcs and Ku70/80 heterodimer) in recruiting other NHEJ repair factors in vivo;2) To test the hypothesis that DNA-PKcs phosphorylation is essential for the NHEJ and also regulates the overall DSB repair machinery by modulating cross talk between NHEJ and HR;3) To test the hypothesis that ATM and DNA-PKcs are coordinated in sensing and repairing the DNA DSBs in mammalian cells;and 4) To determine the functions of DNA-PKcs phosphorylation and kinase activity in transgenic mouse models.
|Mori, Eiichiro; Davis, Anthony J; Hasegawa, Masatoshi et al. (2016) Lysines 3241 and 3260 of DNA-PKcs are important for genomic stability and radioresistance. Biochem Biophys Res Commun 477:235-40|
|Davis, Anthony J; Lee, Kyung-Jong; Chen, David J (2013) The N-terminal region of the DNA-dependent protein kinase catalytic subunit is required for its DNA double-stranded break-mediated activation. J Biol Chem 288:7037-46|
|Wang, Hailong; Shi, Linda Z; Wong, Catherine C L et al. (2013) The interaction of CtIP and Nbs1 connects CDK and ATM to regulate HR-mediated double-strand break repair. PLoS Genet 9:e1003277|
|Shao, Zhengping; Davis, Anthony J; Fattah, Kazi R et al. (2012) Persistently bound Ku at DNA ends attenuates DNA end resection and homologous recombination. DNA Repair (Amst) 11:310-6|
|Pankotai, Tibor; Bonhomme, Celine; Chen, David et al. (2012) DNAPKcs-dependent arrest of RNA polymerase II transcription in the presence of DNA breaks. Nat Struct Mol Biol 19:276-82|
|Chung, Young Min; Park, See-Hyoung; Tsai, Wen-Bin et al. (2012) FOXO3 signalling links ATM to the p53 apoptotic pathway following DNA damage. Nat Commun 3:1000|
|Sun, Jingxin; Lee, Kyung-Jong; Davis, Anthony J et al. (2012) Human Ku70/80 protein blocks exonuclease 1-mediated DNA resection in the presence of human Mre11 or Mre11/Rad50 protein complex. J Biol Chem 287:4936-45|
|Levy-Barda, Adva; Lerenthal, Yaniv; Davis, Anthony J et al. (2011) Involvement of the nuclear proteasome activator PA28? in the cellular response to DNA double-strand breaks. Cell Cycle 10:4300-10|
|Kozlov, Sergei V; Graham, Mark E; Jakob, Burkhard et al. (2011) Autophosphorylation and ATM activation: additional sites add to the complexity. J Biol Chem 286:9107-19|
|Lee, Kyung-Jong; Lin, Yu-Fen; Chou, Han-Yi et al. (2011) Involvement of DNA-dependent protein kinase in normal cell cycle progression through mitosis. J Biol Chem 286:12796-802|
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