Ring-Opened and Tandem DNA Damages

开环和串联 DNA 损伤

• 批准号: 7576741
• 负责人: Ashis K Basu
• 金额: $ 30.1万
• 依托单位: UNIVERSITY OF CONNECTICUT STORRS
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-04-10 至 2011-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7576741
• 关键词: 2' -deoxyadenosine; 8,5' -cyclo-2' -deoxyadenosine; 8,5' -cyclo-2' -deoxyguanosine; 8-Oxo-2' -Deoxyguanosine; 8-hydroxyguanosine; 8-oxo-dA; Abbreviations; Address; Attention; Biological; Bypass; Cells; Circular Dichroism; Classification; DNA; DNA Damage; DNA-Directed DNA Polymerase; Deoxycytidine; Deoxyguanosine; Deoxyuridine; Embryo; Escherichia coli; Etiology; Fapy-dG; Fibroblasts; Genes; Guanine; Human; Imidazole; Ionizing radiation; Knowledge; Lesion; Malignant Neoplasms; Mammalian Cell; Mediating; Modification; Mus; Mutagenesis; Nucleotide Excision Repair; Oligonucleotides; Oncogenes; Organism; Phase; Plasmid Cloning Vector; Plasmids; Play; Process; Proteins; Pyrimidine Dimers; Research; Research Design; Research Personnel; Role; Saccharomyces cerevisiae; Site; TP53 gene; Thymine; Transition Elements; Urea; Viral Vector; base; crosslink; in vivo; insight; melting; molecular dynamics; oxidative DNA damage; programs; repaired; thymine glycol; trend

DESCRIPTION (provided by applicant): We hypothesize that a group of oxidative DNA damages, including 8-oxoguanine, formamidopyrimidines and tandem lesions, play important roles in the etiology of human cancer via the mutagenic and genotoxic effects of these lesions in certain important sequences of cancer genes. In this application, we propose to study the biological effects of a subset of these lesions. We have three specific aims. In aim 1, synthesis and characterization of oligonucleotides containing tandem base lesions in DNA will be carried out. Tandem lesions formed by ionizing radiation, transition metal/H2O2, or other radical- mediated processes include abasic site adjacent to 8-oxoguanine, S- and R-8,5'-cyclo-2'- deoxyguanosine, and intrastrand vicinal guanine-thymine cross-link. We shall determine the structural effects of these damages in DNA by thermal melting, circular dichroism, and NMR. In aim 2, single stranded plasmid or viral vectors containing these lesions will be constructed. Mutagenicity and repair studies of the site-specific lesions will be carried out in Escherichia coli and in mammalian cells. For example, we shall determine how mutagenicity of 8-oxoguanine might be influenced when it is present as part of a tandem DNA damage. We shall investigate if Fapy.dG, the guanine-thymine vicinal cross- link, S- and R-8, 5'-cyclo-2'-deoxyguanosine are mutagenic, genotoxic, or both in mammalian cells. In aim 3, we shall construct duplex plasmid vectors with these lesions, which will be used to examine the extent of lesion bypass by extracts of normal and repair- or replication-altered mammalian cells (e.g., isogenic mouse embryonic fibroblasts and cells that express modulated levels of DNA polymerases encoded by REV1, REV3, RAD18, etc. genes). We hope to contribute toward a deeper insight of the effects of oxidative DNA damages.

描述（由申请人提供）：我们假设一组氧化性 DNA 损伤，包括 8-氧代鸟嘌呤、甲酰胺嘧啶和串联损伤，通过这些损伤在某些重要序列中的诱变和基因毒性作用，在人类癌症的病因学中发挥重要作用。癌症基因。在此应用中，我们建议研究这些病变子集的生物学效应。我们有三个具体目标。在目标 1 中，将进行含有 DNA 串联碱基损伤的寡核苷酸的合成和表征。由电离辐射、过渡金属/H2O2或其他自由基介导的过程形成的串联损伤包括邻近8-氧代鸟嘌呤、S-和R-8,5'-环-2'-脱氧鸟苷以及链内邻位鸟嘌呤-胸腺嘧啶的脱碱基位点交叉链接。我们将通过热熔解、圆二色性和核磁共振来确定 DNA 中这些损伤的结构效应。在目标2中，将构建含有这些损伤的单链质粒或病毒载体。将在大肠杆菌和哺乳动物细胞中进行特定位点损伤的致突变性和修复研究。例如，我们将确定当 8-氧代鸟嘌呤作为串联 DNA 损伤的一部分存在时，其致突变性可能受到怎样的影响。我们将研究 Fapy.dG、鸟嘌呤-胸腺嘧啶邻位交联、S-和 R-8、5'-环-2'-脱氧鸟苷在哺乳动物细胞中是否具有诱变性、遗传毒性或两者兼而有之。在目标 3 中，我们将构建具有这些病变的双链体质粒载体，该载体将用于通过正常和修复或复制改变的哺乳动物细胞（例如，同基因小鼠胚胎成纤维细胞和表达调节的细胞）的提取物来检查病变旁路的程度。 REV1、REV3、RAD18 等基因编码的 DNA 聚合酶的水平）。我们希望有助于更深入地了解 DNA 氧化损伤的影响。