Nonhomologous end joining (NHEJ) is a major pathway for resolving chromosome double strand breaks. NHEJ deficiency results in a wide variety of phenotypes including premature cellular senescence, immunodeficiency, defective neurogenesis, dwarfism, accelerated aging, and cancer predisposition. Notably, NHEJ is mostly dispensable in resolving the "clean" chromosome breaks used in many experimental models. Why, then, are the phenotypes of NHEJ deficiency so severe? We will address here if NHEJ's pivotal role in maintaining genome stability is at least in part due to a unique ability to flexibly cope with the complex end structures expected in biologically relevant contexts. Of particular interest to this proposal, broken ends generated by ionizing radiation, radiomimetic drugs, or chronic oxidative stress possess associated damage to flanking DNA, including oxidized and missing bases. The ability of ionizing radiation and radiomimetic drugs to introduce such complex damage is a major factor in the biological impact of these agents, and thus critical in the effectiveness of these agents in tumor therapy.
In Aim 1, we will determine how associated radiomimetic damage effects how NHEJ joins ends, both in vitro and in cells. Ku is a key factor in repair of breaks by NHEJ.
In Aim 2 we will address the extent to which Ku's recently described ability to excise damaged nucleotides from ends contributes to its role in NHEJ.
In Aim 3 we will determine how NHEJ regulates the fidelity of the ligation step, and what impact regulation of fidelity has on resolution of complex end structures.

Public Health Relevance

This work represents a systematic study addressing how our cells resolve chromosome breaks caused by chronic stress or acute radiation exposure. Our work will provide insights into aging and how radiation kills cells, and will be used to generate safer and more effective ways to use radiation as a tool for therapy.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project (R01)
Project #
5R01CA084442-13
Application #
8258788
Study Section
Cancer Etiology Study Section (CE)
Program Officer
Pelroy, Richard
Project Start
2000-01-04
Project End
2014-04-30
Budget Start
2012-05-01
Budget End
2013-04-30
Support Year
13
Fiscal Year
2012
Total Cost
$251,202
Indirect Cost
$80,683
Name
University of North Carolina Chapel Hill
Department
Biochemistry
Type
Schools of Medicine
DUNS #
608195277
City
Chapel Hill
State
NC
Country
United States
Zip Code
27599
Kusumoto-Matsuo, Rika; Ghosh, Deblina; Karmakar, Parimal et al. (2014) Serines 440 and 467 in the Werner syndrome protein are phosphorylated by DNA-PK and affects its dynamics in response to DNA double strand breaks. Aging (Albany NY) 6:70-81
Williams, Gareth J; Hammel, Michal; Radhakrishnan, Sarvan Kumar et al. (2014) Structural insights into NHEJ: building up an integrated picture of the dynamic DSB repair super complex, one component and interaction at a time. DNA Repair (Amst) 17:110-20
Waters, Crystal A; Strande, Natasha T; Wyatt, David W et al. (2014) Nonhomologous end joining: a good solution for bad ends. DNA Repair (Amst) 17:39-51
Strande, Natasha T; Carvajal-Garcia, Juan; Hallett, Ryan A et al. (2014) Requirements for 5'dRP/AP lyase activity in Ku. Nucleic Acids Res 42:11136-43
Waters, Crystal A; Strande, Natasha T; Pryor, John M et al. (2014) The fidelity of the ligation step determines how ends are resolved during nonhomologous end joining. Nat Commun 5:4286
Strande, Natasha; Roberts, Steven A; Oh, Sehyun et al. (2012) Specificity of the dRP/AP lyase of Ku promotes nonhomologous end joining (NHEJ) fidelity at damaged ends. J Biol Chem 287:13686-93
Ramsden, Dale A (2011) Polymerases in nonhomologous end joining: building a bridge over broken chromosomes. Antioxid Redox Signal 14:2509-19
Roberts, Steven A; Strande, Natasha; Burkhalter, Martin D et al. (2010) Ku is a 5'-dRP/AP lyase that excises nucleotide damage near broken ends. Nature 464:1214-7
Kusumoto-Matsuo, Rika; Opresko, Patricia L; Ramsden, Dale et al. (2010) Cooperation of DNA-PKcs and WRN helicase in the maintenance of telomeric D-loops. Aging (Albany NY) 2:274-84
Ramsden, Dale A; Weed, Brett D; Reddy, Yeturu V R (2010) V(D)J recombination: Born to be wild. Semin Cancer Biol 20:254-60

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