A growing body of evidence suggests that endogenous damage to DNA may contribute significantly to genomic instability, human disease and aging. An understanding of the formation and persistence of endogenous DNA damage is also required in order to assess the impact of DNA damage induced by exogenous mutagens and carcinogens.The purpose of the work described in this proposal is to investigate several components of endogenous DNA damage. Endogenous damage to DNA may result from either hydrolysis or oxidation of DNA and its components. Among the purines, hydrolysis resulting in the abasic sites and oxidation to form 8-oxopurines have dominated the literature. Purines may also be damaged by imidazole ring-opening generating formamidopyrimidine (FAPY) derivatives via both hydrolysis and oxidation. Hydrolytic formation of FAPY derivatives has received minimal attention in the literature. The kinetics of purine hydrolysis must be understood in order to assess accurately the significance of FAPY derivatives.The preliminary results from Dr. Sowers' laboratory indicate that current and often reported measurements of FAPY derivatives may yield extreme underestimates of the actual levels. Similarly, hydrolysis of the oxopurines may be biologically significant, and can lead to erroneous quantitation of oxidation damaged products. Oxidation of thymidine generating 5- hydroxymethyl-2'-deoxyuridine (HMdU) is a frequent DNA lesion. HMdU is controversial in terms of both amount formed and biological significance. Dr. Sowers' laboratory has identified chemical characteristics of HMdU which may explain the divergent quantitative reports. The investigators propose a series of experiments which should shed light on the potential biological impact of HMdU formation in DNA. In parallel with the oxidation of the thymine methyl group, they present preliminary data showing that the methyl group of 5-methylcytosine is also a target for oxidative damage. The oxidative damage to 5- methylcytosine is relatively unexplored. Because of the pivotal role of cytosine methylation in gene control, oxidation of 5mC may be part of an unrecognized mechanism for inappropriate activation of oncogenes and latent viral genes.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM050351-05
Application #
6179671
Study Section
Special Emphasis Panel (ZRG2-MEP (01))
Program Officer
Wolfe, Paul B
Project Start
1994-01-01
Project End
2001-03-31
Budget Start
2000-04-01
Budget End
2001-03-31
Support Year
5
Fiscal Year
2000
Total Cost
$263,720
Indirect Cost
Name
City of Hope/Beckman Research Institute
Department
Type
DUNS #
City
Duarte
State
CA
Country
United States
Zip Code
91010
Theruvathu, Jacob A; Darwanto, Agus; Hsu, Chia Wei et al. (2014) The effect of Pot1 binding on the repair of thymine analogs in a telomeric DNA sequence. Nucleic Acids Res 42:9063-73
Theruvathu, Jacob A; Yin, Y Whitney; Pettitt, B Montgomery et al. (2013) Comparison of the structural and dynamic effects of 5-methylcytosine and 5-chlorocytosine in a CpG dinucleotide sequence. Biochemistry 52:8590-8
Theruvathu, Jacob A; Kim, Cherine H; Darwanto, Agus et al. (2009) pH-Dependent configurations of a 5-chlorouracil-guanine base pair. Biochemistry 48:11312-8
Darwanto, Agus; Farrel, Alvin; Rogstad, Daniel K et al. (2009) Characterization of DNA glycosylase activity by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. Anal Biochem 394:13-23
Liu, Pingfang; Theruvathu, Jacob A; Darwanto, Agus et al. (2008) Mechanisms of base selection by the Escherichia coli mispaired uracil glycosylase. J Biol Chem 283:8829-36
Valinluck, Victoria; Sowers, Lawrence C (2007) Inflammation-mediated cytosine damage: a mechanistic link between inflammation and the epigenetic alterations in human cancers. Cancer Res 67:5583-6
Valinluck, Victoria; Sowers, Lawrence C (2007) Endogenous cytosine damage products alter the site selectivity of human DNA maintenance methyltransferase DNMT1. Cancer Res 67:946-50
Rogstad, Daniel K; Darwanto, Agus; Herring, Jason L et al. (2007) Measurement of the incorporation and repair of exogenous 5-hydroxymethyl-2'-deoxyuridine in human cells in culture using gas chromatography-negative chemical ionization-mass spectrometry. Chem Res Toxicol 20:1787-96
Valinluck, Victoria; Liu, Pingfang; Kang Jr, Joseph I et al. (2005) 5-halogenated pyrimidine lesions within a CpG sequence context mimic 5-methylcytosine by enhancing the binding of the methyl-CpG-binding domain of methyl-CpG-binding protein 2 (MeCP2). Nucleic Acids Res 33:3057-64
Valinluck, Victoria; Tsai, Hsin-Hao; Rogstad, Daniel K et al. (2004) Oxidative damage to methyl-CpG sequences inhibits the binding of the methyl-CpG binding domain (MBD) of methyl-CpG binding protein 2 (MeCP2). Nucleic Acids Res 32:4100-8

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