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）：Translesion合成（TLS）DNA聚合酶（POLS）有助于确保通过通过DNA病变促进复制来确保复制叉的持续进展。在拟议的研究中，我们将通过各种由环境污染物和致癌物以及细胞氧化损伤诱导的多种DNA病变来确定许多人类TLS POL在促进复制中的作用。特别是，我们将检验以下假设：通过人类细胞中的DNA病变复制是通过两种不同模式发生的，在哪种pol，？，k，rev1和？主要介导无错误的TL，并以高度专业的方式作用于DNA病变，而POL？以更普遍和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的更普遍和容易产生错误的作用？源自其插入嘌呤核苷酸（NT）的能力，优先是通过“蛋白质 - 板”指导机制的A，相反的DNA病变。为了通过人类的DNA病变阐明无错误和诱变复制的遗传碱基，我们将进行以下研究。在AIM 1中，我们将研究人和小鼠细胞中各种TLS POLS对无误差与诱变病变旁路的贡献。 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致癌物，1，N2-丙烷-2'-脱氧瓜烷（PDG），这是一种由脂质过氧化产生的环链形式，它也是一种无处不在的环境污染物，以及N2-DG的N2-DG加合物，将环境癌（+）抗抗苯甲酸蛋白酶抗蛋白毒性加入。在AIM 2中，将进行生化研究以检查POL的水平吗？在合成相对的DNA（6-4）TT光产物，8-oxog，EDA，PDG和N2-DG BPDE中。通过稳态动力学分析，我们将确定POL的催化效率？为了插入与DNA病变相对的核苷酸并进行随后的延伸反应，并且将进行生化研究以检验POL的假设？通过蛋白质模板指导的机制插入与DNA病变相对的嘌呤NT。 我们在此提出的遗传和生化研究与描述TLS POL在复制过程中复制过程中促进TLS POL的作用高度相关，并为对环境和细胞DNA在人类细胞中诱导的诱变和细胞DNA诱导的遗传学生成的遗传生成的遗传生成具有全面的理解。我们提出的提议主要通过POLS？，？，K，REV1和？这些POL在癌症抑制中的作用是否会预测，而POL的TLS诱变模式？将预测该POL在增强基因组不稳定性和癌变中的作用。 公共卫生相关性：人类细胞中的DNA病变是由于细胞氧化损伤以及暴露于化学和环境致癌物而产生的。在人类细胞复制过程中促进无错性病变旁路中各种DNA聚合酶在人类细胞复制过程中的作用的确定对于提供对由环境和细胞DNA损害药物引起的诱变诱导的诱变和癌变的遗传基础的全面理解。