We will determine whether base excision repair of oxidatively damaged DNA bases occurs preferentially in actively transcribed genes. The repair of 5-hydroxymethyluracil (HmUra), which is repaired by HmUra-DNA glycosylase and does not block replication, will be compared to that of thymine glycol and related lesions which are repaired by E. coli endonuclease III and do block replication. HmUra-DNA glycosylase is unique among repair enzymes in being absent from bacteria and yeast. We have proposed that its phylogenetic distribution reflects its major function which is the repair of 5-methylcytosine residues in DNA. In contrast, endonuclease III-like activities are phylogenetically conserved indicating that the lesions repaired by the enzymes are formed in all species. Among these lesions are UV induced pyrimidine hydrates. The hydration of cytosine leads to irreversible deamination to uracil hydrate which, if unrepaired, leads to point mutations. This reaction scheme may also occur for 5-methylcytosine residues leading to irreversible deamination to thymine hydrate. We will study the formation and stability of pyrimidine photohydrates in oligonucleotides of high specific radioactivity using E. coli endonuclease III as a probe. We will use these oligonucleotides as substrates for the purification of mammalian endonuclease III-like activities to a purity sufficient to obtain protein sequence and molecular cloning. The results of these experiments will add to our knowledge of the formation and repair of oxidative and photochemical damage to DNA and possibly elucidate the relationship of such damage to the etiology of actinic and spontaneous cancer.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project (R01)
Project #
5R01CA049869-05
Application #
3194187
Study Section
Metabolic Pathology Study Section (MEP)
Project Start
1989-04-15
Project End
1995-06-30
Budget Start
1993-07-01
Budget End
1994-06-30
Support Year
5
Fiscal Year
1993
Total Cost
Indirect Cost
Name
New York University
Department
Type
Schools of Medicine
DUNS #
004514360
City
New York
State
NY
Country
United States
Zip Code
10012
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Marenstein, D R; Ocampo, M T; Chan, M K et al. (2001) Stimulation of human endonuclease III by Y box-binding protein 1 (DNA-binding protein B). Interaction between a base excision repair enzyme and a transcription factor. J Biol Chem 276:21242-9
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Zuo, S; Boorstein, R J; Teebor, G W (1995) Oxidative damage to 5-methylcytosine in DNA. Nucleic Acids Res 23:3239-43
O'Donnell, R E; Boorstein, R J; Cunningham, R P et al. (1994) Effect of pH and temperature on the stability of UV-induced repairable pyrimidine hydrates in DNA. Biochemistry 33:9875-80
Lustig, M J; Cadet, J; Boorstein, R J et al. (1992) Synthesis of the diastereomers of thymidine glycol, determination of concentrations and rates of interconversion of their cis-trans epimers at equilibrium and demonstration of differential alkali lability within DNA. Nucleic Acids Res 20:4839-45
Levy, D D; Teebor, G W (1991) Site directed substitution of 5-hydroxymethyluracil for thymine in replicating phi X-174am3 DNA via synthesis of 5-hydroxymethyl-2'-deoxyuridine-5'-triphosphate. Nucleic Acids Res 19:3337-43

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