DNA damage causes mutation or cell death and is a contributing factor to carcinogenesis. Our laboratory has focused on DNA damage by electrophilic species generated endogenously by oxidative stress or inflammation. We are particularly interested in the lipid oxidation product, malondialdehyde (MDA), and the DNA oxidation products, base propenals. Previous work from our laboratory has provided a comprehensive inventory of adducts formed on reaction of MDA/base propenals with deoxynucleosides and DNA;demonstrated that the major adduct to deoxyguanosine (MldG) is mutagenic in bacterial and mammalian cells;revealed that MldG is repaired by nucleotide excision repair;and provided rigorous anal3^ical evidence that MldG is present endogenously in the human genome. In the most recent grant period, we have provided a detailed kinetic and structural analysis of the induction of mutations by MldG by Y-family translesion polymerases including a crystal structure of an MldG-containing template-primer bound in the active site of Dpo4;discovered that the major adduct of MDA/base propenals to deoxyadenosine, which is called OPdA, reacts with amino acids to form stable cross-links to DNA;and reported that MldG is present in the urine of healthy human subjects along with a single major metabolite, 6-oxo-MldG. These discoveries place us in an outstanding position to explore the chemical biology of DNA damage by endogenous products of oxidative stress and inflammation.
Our specific aims for the next grant period are to (1) Determine the structural and functional consequences of the interaction of template-primers containing the endogenous adducts, MldG, OPdA, and heptanone-etheno-dG with Y-family DNA polymerases;(2) Explore the chemistry of DNA-DNA and DNA-protein cross-linking mediated by the endogenous adducts, OPdA and OPdC;and (3) Develop highly sensitive and specific analytical methods for quantification of MldG and 6-oxo-MldG that will allow simultaneous monitoring of both lesions in human populations and animal models.

Public Health Relevance

Cancer is caused by mutations to normal genes that regulate processes such as growth, survival, and invasion. Our laboratory discovered that DNA damage, which is a major cause of genetic mutations, can occur from normal metabolic processes such as inflammation. Since the relationship between chronic inflammation and cancer has been known for over a hundred years, our research should help to define the molecular basis for this major cause of human cancer.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Method to Extend Research in Time (MERIT) Award (R37)
Project #
5R37CA087819-15
Application #
8677568
Study Section
Special Emphasis Panel (NSS)
Program Officer
Okano, Paul
Project Start
2000-06-01
Project End
2015-05-31
Budget Start
2014-06-01
Budget End
2015-05-31
Support Year
15
Fiscal Year
2014
Total Cost
$408,918
Indirect Cost
$140,184
Name
Vanderbilt University Medical Center
Department
Biochemistry
Type
Schools of Medicine
DUNS #
004413456
City
Nashville
State
TN
Country
United States
Zip Code
37212
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Shuck, Sarah C; Wauchope, Orrette R; Rose, Kristie L et al. (2014) Protein modification by adenine propenal. Chem Res Toxicol 27:1732-42
Beavers, William N; Serwa, Remigiusz; Shimozu, Yuki et al. (2014) ?-Alkynyl lipid surrogates for polyunsaturated fatty acids: free radical and enzymatic oxidations. J Am Chem Soc 136:11529-39
Shuck, Sarah C; Rose, Kristie L; Marnett, Lawrence J (2014) Mass spectrometric methods for the analysis of nucleoside-protein cross-links: application to oxopropenyl-deoxyadenosine. Chem Res Toxicol 27:136-46
Akingbade, Dapo; Kingsley, Philip J; Shuck, Sarah C et al. (2012) Selection of monoclonal antibodies against 6-oxo-M(1)dG and their use in an LC-MS/MS assay for the presence of 6-oxo-M(1)dG in vivo. Chem Res Toxicol 25:454-61
Maddukuri, Leena; Eoff, Robert L; Choi, Jeong-Yun et al. (2010) In vitro bypass of the major malondialdehyde- and base propenal-derived DNA adduct by human Y-family DNA polymerases ?, ?, and Rev1. Biochemistry 49:8415-24
Eoff, Robert L; McGrath, Colleen E; Maddukuri, Leena et al. (2010) Selective modulation of DNA polymerase activity by fixed-conformation nucleoside analogues. Angew Chem Int Ed Engl 49:7481-5
Cline, Susan D; Lodeiro, M Fernanda; Marnett, Lawrence J et al. (2010) Arrest of human mitochondrial RNA polymerase transcription by the biological aldehyde adduct of DNA, M1dG. Nucleic Acids Res 38:7546-57
Uddin, Md Jashim; Schulte, Michael I; Maddukuri, Leena et al. (2010) Semisynthesis of 6-chloropurine-2'-deoxyriboside 5'-dimethoxytrityl 3'-(2-cyanoethyl-N,N-diisopropylamino)phosphoramidite and its use in the synthesis of fluorescently labeled oligonucleotides. Nucleosides Nucleotides Nucleic Acids 29:831-40
Knutson, Charles G; Rubinson, Emily H; Akingbade, Dapo et al. (2009) Oxidation and glycolytic cleavage of etheno and propano DNA base adducts. Biochemistry 48:800-9

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