Genetic control of replication through DNA lesions in humans, and carcinogenesis

通过人类 DNA 损伤对复制进行遗传控制以及致癌作用

• 批准号: 8216401
• 负责人: SATYA PRAKASH
• 金额: $ 34.43万
• 依托单位: UNIVERSITY OF TEXAS MED BR GALVESTON
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-01-27 至 2016-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8216401
• 关键词: 7,8-dihydro-8-oxoguanine; Acrolein; Adenine; Benzo(a)pyrene; Biochemical; Biochemical Reaction; Biological; Bypass; Cells; DNA; DNA Damage; DNA biosynthesis; DNA lesion; DNA-Directed DNA Polymerase; Deoxyguanosine; Ensure; Environmental Carcinogens; Environmental Pollutants; Epoxy Compounds; Exposure to; Family; Free Radicals; Frequencies; Genes; Genetic; Genomic Instability; Glycols; Guanine; Human; Kinetics; Lesion; Lipid Peroxidation; Malignant Neoplasms; Mediating; Mus; Mutagenesis; Mutation; Nucleotides; Pathway interactions; Plasmids; Play; Polymerase; Proteins; Purine Nucleotides; Relative (related person); Role; Scheme; Simian virus 40; Small Interfering RNA; System; Testing; adduct; base; carcinogenesis; chemical carcinogen; in vivo; oxidative damage; ultraviolet irradiation

DESCRIPTION (provided by applicant): Translesion synthesis (TLS) DNA polymerases (Pols) help ensure the continued progression of the replication fork by promoting replication through DNA lesions. In the proposed studies, we will determine the roles of a number of human TLS Pols in promoting replication through a variety of DNA lesions induced by environmental pollutants and carcinogens and by cellular oxidative damage. In particular, we will test the hypothesis that replication through DNA lesions in human cells occurs via two distinct modes in which Pols, ?, ?, k, Rev1, and ? mediate predominantly error-free TLS and act in a highly specialized manner dependent upon the DNA lesion, whereas Pol? performs lesion bypass in a more generalized and error-prone manner. Further, we will test the hypothesis that the more generalized and error-prone role of Pol? emanates from its ability to insert a purine nucleotide (nt), preferentially an A, opposite DNA lesions via a "protein-template"-directed mechanism. To elucidate the genetic bases of error-free and mutagenic replication through DNA lesions in humans, we will carry out the following studies. In Aim 1, we will examine the contributions of various TLS Pols to error-free vs. mutagenic lesion bypass in human and mouse cells. The lesions to be studied include (6- 4) photoproduct induced by UV irradiation, 7,8-dihydro-8-oxogunaine (8-oxoG) generated from free-radical attack on guanine in DNA, 1,N6-ethenodeoxyadenosine (edA) generated from interaction of adenine with products of lipid peroxidation resulting from cellular oxidative damage and from exposure to chemical carcinogens, 1,N2-propano-2'-deoxyguanosine (PdG), a ring-closed form of acrolein generated from lipid peroxidation and which also is a ubiquitous environmental pollutant, and N2-dG adduct of the environmental carcinogen (+) anti-benzo[a]pyrene diol expoxide (BPDE). In Aim 2, biochemical studies will be done to examine the proficiency of Pol? in synthesizing DNA opposite (6-4) TT photoproduct, 8-oxoG, edA, PdG, and N2-dG BPDE. By steady-state kinetic analyses we will determine the catalytic efficiency of Pol? for inserting nucleotides opposite the DNA lesion and for carrying out the subsequent extension reaction, and biochemical studies will be done to test the hypothesis that Pol? inserts a purine nt opposite DNA lesions via a protein- template-directed mechanism. The genetic and biochemical studies we propose here are highly relevant for delineating the roles of TLS Pols in promoting error-free vs. mutagenic lesion bypass during replication, and for providing a comprehensive understanding of the genetic bases of mutagenesis and carcinogenesis induced by environmental and cellular DNA damaging agents in human cells. Our proposal for a predominantly error-free mode of TLS by Pols ?, ?, k, Rev1, and ? would predict a role for these Pols in cancer suppression, whereas a mutagenic mode of TLS by Pol? would predict a role for this Pol in enhancing genomic instability and carcinogenesis. PUBLIC HEALTH RELEVANCE: DNA lesions are generated in human cells from cellular oxidative damage and from exposure to chemical and environmental carcinogens. The determination of roles of various DNA polymerases in promoting error-free vs. mutagenic lesion bypass during replication in human cells is important for providing a comprehensive understanding of the genetic bases of mutagenesis and carcinogenesis induced by environmental and cellular DNA damaging agents.

描述（由申请人提供）：跨损伤合成（TLS）DNA聚合酶（Pols）通过DNA损伤促进复制，有助于确保复制叉的持续进展。在拟议的研究中，我们将确定许多人类 TLS Pols 在通过环境污染物和致癌物以及细胞氧化损伤引起的各种 DNA 损伤促进复制中的作用。特别是，我们将测试这样的假设：人类细胞中通过 DNA 损伤进行的复制通过两种不同的模式发生，其中 Pols、?、?、k、Rev1 和 ?主要介导无错误的 TLS，并根据 DNA 损伤以高度专业化的方式起作用，而 Pol？以更通用且更容易出错的方式执行病变旁路。此外，我们将检验以下假设：Pol 的作用更普遍且更容易出错。源自其通过“蛋白质模板”导向机制插入嘌呤核苷酸（nt）（优先是 A）相对 DNA 损伤的能力。为了阐明通过人类DNA损伤进行无差错和诱变复制的遗传基础，我们将进行以下研究。在目标 1 中，我们将检查各种 TLS Pol 对人类和小鼠细胞中无错与诱变病变旁路的贡献。要研究的损伤包括 (6- 4) 紫外线照射诱导的光产物、DNA 中鸟嘌呤自由基攻击产生的 7,8-二氢-8-氧代鸟嘌呤 (8-oxoG)、1,N6-乙脱氧腺苷 (edA)由腺嘌呤与细胞氧化损伤和接触化学致癌物引起的脂质过氧化产物相互作用而产生， 1,N2-丙-2'-脱氧鸟苷（PdG），脂质过氧化产生的丙烯醛的闭环形式，也是一种普遍存在的环境污染物，是环境致癌物（+）抗苯并的N2-dG加合物。 a]芘二醇环氧化物（BPDE）。在目标 2 中，将进行生化研究来检查 Pol? 的熟练程度。合成与 (6-4) TT 光产物、8-oxoG、edA、PdG 和 N2-dG BPDE 相反的 DNA。通过稳态动力学分析，我们将确定 Pol? 的催化效率。用于在DNA损伤对面插入核苷酸并进行随后的延伸反应，并且将进行生化研究来检验Pol？通过蛋白质模板导向机制在 DNA 损伤对面插入嘌呤 nt。 我们在此提出的遗传和生化研究与描述 TLS Pols 在复制过程中促进无错与诱变病变旁路的作用高度相关，并提供对环境和细胞诱导的诱变和致癌作用的遗传基础的全面理解。人体细胞中的 DNA 损伤剂。我们对 Pols ?、?、k、Rev1 和 ? 提出的主要无错误 TLS 模式的建议将预测这些 Pols 在癌症抑制中的作用，而 Pol? 的 TLS 的诱变模式？将预测该 Pol 在增强基因组不稳定性和致癌作用中的作用。 公共健康相关性：DNA 损伤是由于细胞氧化损伤以及接触化学和环境致癌物而在人体细胞中产生的。确定各种 DNA 聚合酶在促进人类细胞复制过程中无错误与诱变病变绕过中的作用对于全面了解环境和细胞 DNA 损伤剂诱导的诱变和致癌的遗传基础非常重要。