DNA double stranded-breaks (DSBs) are thought to be the most dangerous DNA lesions that threaten genomic integrity. In vertebrates, the classical non-homologous end joining pathway (c-NHEJ) joins most DSBs in all cell types and at all stages of the cell cycle. Not surprisingly, intact c-NHEJ is essential for human life. Disruption o c-NHEJ in somatic cells results in genomic instability and is strongly associated with tumorigenesis. Paradoxically, strategies for disrupting c-NHEJ, may have several potentially important therapeutic applications. Thus, a solid understanding of this pathway is important to human health. Although c-NHEJ has been studied for decades, more basic research is warranted because our knowledge of c-NHEJ is not only incomplete, but may also be inaccurate. The current research focuses on three areas. The first will ascertain mechanistic details of how phosphorylations mediated by the DNA dependent protein kinase (DNA-PK) regulate DNA repair. DNA-PK is a large protein kinase complex that initiates DNA repair. It targets (by protein phosphorylation) all of the c-NHEJ factors including itself. This autophosphorylation is essential for repair and is functionally complex. The second area of research will focus on emerging data demonstrating how DNA-PK affects other DNA repair pathways. c-NHEJ should not join certain types of DNA damage;one example of damage that c-NHEJ should preferably avoid is the damage that results during DNA replication when a replication fork collapses.
The second aim will determine whether DNA-PK promotes cell survival during replication stress at the expense of genome integrity.
The third aim follows up on recent novel findings showing how XRCC4 and XLF [two factors previously ascribed to c-NHEJ's ligation complex, and thus factors that function late in c-NHEJ] form filamentous structures that bridge DNA in vitro, and that may function in living cells to stabilize DNA ends prior to repair. This suggests an additional, early (in c-NHEJ) role for XRCC4/XLF filaments. This would be a significant shift from current dogma.

Public Health Relevance

DNA (the blue print for all life) is very sensitive and can be damaged in many ways;the pathway that repairs double stranded-breaks is essential to human life. Disruption of this pathway prior to birth results in Severe Combined Immunodeficiency (SCID), and in most cases, severe brain defects;whereas disruption of this pathway after birth can lead to cancer. The current research will ascertain mechanistic details of this DNA repair pathway;this basic research is warranted because our knowledge of this type of DNA repair is not only incomplete, but may also be inaccurate.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Research Project (R01)
Project #
5R01AI048758-15
Application #
8662140
Study Section
Cellular and Molecular Immunology - A Study Section (CMIA)
Program Officer
Nasseri, M Faraz
Project Start
2000-03-15
Project End
2017-04-30
Budget Start
2014-05-01
Budget End
2015-04-30
Support Year
15
Fiscal Year
2014
Total Cost
$377,509
Indirect Cost
$127,509
Name
Michigan State University
Department
Pathology
Type
Schools of Veterinary Medicine
DUNS #
193247145
City
East Lansing
State
MI
Country
United States
Zip Code
48824
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Neal, Jessica A; Sugiman-Marangos, Seiji; VanderVere-Carozza, Pamela et al. (2014) Unraveling the complexities of DNA-dependent protein kinase autophosphorylation. Mol Cell Biol 34:2162-75
Woodbine, Lisa; Neal, Jessica A; Sasi, Nanda-Kumar et al. (2013) PRKDC mutations in a SCID patient with profound neurological abnormalities. J Clin Invest 123:2969-80
Meek, Katheryn; Lees-Miller, Susan P; Modesti, Mauro (2012) N-terminal constraint activates the catalytic subunit of the DNA-dependent protein kinase in the absence of DNA or Ku. Nucleic Acids Res 40:2964-73
Roy, Sunetra; Andres, Sara N; Vergnes, Alexandra et al. (2012) XRCC4's interaction with XLF is required for coding (but not signal) end joining. Nucleic Acids Res 40:1684-94
Neal, Jessica A; Dang, Van; Douglas, Pauline et al. (2011) Inhibition of homologous recombination by DNA-dependent protein kinase requires kinase activity, is titratable, and is modulated by autophosphorylation. Mol Cell Biol 31:1719-33
Neal, Jessica A; Meek, Katheryn (2011) Choosing the right path: does DNA-PK help make the decision? Mutat Res 711:73-86
Shibata, Atsushi; Conrad, Sandro; Birraux, Julie et al. (2011) Factors determining DNA double-strand break repair pathway choice in G2 phase. EMBO J 30:1079-92
Muller, Tina A; Meek, Katheryn; Hausinger, Robert P (2010) Human AlkB homologue 1 (ABH1) exhibits DNA lyase activity at abasic sites. DNA Repair (Amst) 9:58-65
Cui, Xiaoping; Meek, Katheryn (2007) Linking double-stranded DNA breaks to the recombination activating gene complex directs repair to the nonhomologous end-joining pathway. Proc Natl Acad Sci U S A 104:17046-51

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