Role of human DNA polymerase kappa in replicative bypass of DNA lesions

人类 DNA 聚合酶 kappa 在 DNA 损伤复制旁路中的作用

• 批准号: 8464657
• 负责人: SATYA PRAKASH
• 金额: $ 42.29万
• 依托单位: UNIVERSITY OF TEXAS MED BR GALVESTON
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-07-01 至 2016-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8464657
• 关键词: 4 hydroxynonenal; 7,8-Dihydro-7,8-dihydroxybenzo(a)pyrene 9,10-oxide; 7,8-dihydro-8-oxoguanine; Active Sites; Address; Adverse effects; Affect; Aldehydes; Aromatic Polycyclic Hydrocarbons; Automobiles; Benzo(a)pyrene; Binding; Biochemical; Biochemical Genetics; Bypass; Cancer Biology; Cancer Etiology; Cells; Complex; DNA; DNA Adducts; DNA biosynthesis; DNA lesion; DNA-Directed DNA Polymerase; Deoxyguanosine; Ensure; Environmental Carcinogens; Environmental Pollutants; Epoxy Compounds; Exposure to; Family; Free Radicals; Genetic; Genome Stability; Glycols; Guanine; Human; Hydroxyl Radical; Incidence; Lesion; Lipid Peroxidation; Lipids; Malignant Neoplasms; Mammalian Cell; Mediating; Membrane; Minor Groove; Molecular; Mutation; Nucleotides; Plasmids; Polymerase; Process; Reaction; Role; Simian virus 40; Solvents; Specificity; Structure; System; Testing; Time; UV induced; adduct; base; benzo(a)pyrene 7,8-diol-9,10-epoxide-N2-deoxyguanosine; carcinogenesis; chemical carcinogen; dimer; human DNA; oxidative DNA damage; oxidative damage; public health relevance; thymine glycol

DESCRIPTION (provided by applicant): Translesion synthesis (TLS) DNA polymerases (Pols) promote replication through DNA lesions. Humans possess four TLS Pols that belong to the Y-family, Pols ?, ?, ?, and Rev1, and another Pol, Pol?, that belongs to the B-family. These TLS Pols employ highly specialized mechanisms for replicating through DNA lesions. Of these, Pol? is adept at extending opposite from bulky N2-dG minor groove lesions. Pol? can also carry out TLS opposite other types of DNA lesions; in that case, however, the lesion bypass ability is not limited to Pol?, as other TLS Pols can also function in their bypass. To elucidate the role of Pol? in lesion bypass in human cells, we will use a combined biochemical, genetic, and structural approach. In Aim 1, the role of two unique structural features of Pol?, (i) the N-clasp which allows Pol? to encircle DNA, and (ii) the openness of its active site towards the minor groove at the template-primer junction, will be analyzed by determining the effects of mutations on the extension reaction from N2-dG minor groove adducts and other types of DNA lesions as well. In Aim 2, the role of Pol? in promoting replication through different types of DNA lesions in human cells will be analyzed using a newly devised SV40-based plasmid system. Among the DNA lesions to be studied are 8- oxoguanine (8-oxoG) and thymine glycol (TG) that result from cellular oxidative DNA damage; the ring-opened N2-(3-hydroxyl propyl-2'-deoxygunaosine [(r)-3HOPdG] and the bulky trans-4-hydroxy-2-non-enal- deoxyguanosine (HNE-dG) adducts that result from the reaction of N2-dG with aldehydes or enals generated from free radical attack on lipids in membranes; and the multi-cyclic benzo[a]pyrene 7,8-diol 9,10-epoxide (BPDE) N2-dG adduct that results from exposure to environmental pollutants and carcinogens. To gain a deeper understanding of how Pol? actually performs TLS opposite these DNA lesions, in Aim 3, we will determine crystal structures of Pol? with 8-oxoG, TG, and cis-syn TT dimer, as well as with the N2-dG minor groove adducts of (r)-3HOPdG, HNE, and BPDE. The proficient ability of Pol? for extending from the C inserted opposite the N2-dG adducts by another DNA Pol, such as ? or Rev1, would ensure error-free replication through such minor groove DNA adducts. Since a large variety of N2-dG adducts are formed in human cells from cellular oxidative reactions and from exposure to chemical and environmental carcinogens, Pol? will have a major impact on genome stability by keeping the rate of mutations low, reducing thereby the incidence of carcinogenesis in humans. The proposed studies are highly relevant for cancer biology and etiology as they will reveal how human cells minimize the mutagenic and carcinogenic potential of DNA lesions.

描述（由申请人提供）：Translesion合成（TLS）DNA聚合酶（POLS）通过DNA病变促进复制。人类拥有四个属于Y家族的TLS pol，Pols？，？，？和rev1，以及另一个属于B家族的pol？这些TLS POL采用高度专业的机制通过DNA病变进行复制。其中，Pol？善于与笨重的N2-DG小凹槽病变相对。 pol？还可以执行与其他类型的DNA病变相对的TL；但是，在这种情况下，病变旁路能力不限于POL？，因为其他TLS POLS也可以在其旁路中起作用。阐明POL的作用？在人类细胞的病变旁路中，我们将使用合并的生化，遗传和结构方法。在AIM 1中，Pol的两个独特结构特征的作用？为了环绕DNA，（ii）将通过确定突变对N2-DG小凹槽加合物和其他类型的DNA病变的扩展反应的影响来分析其活跃位点向模板 - 播种机连接处的次要凹槽的开放性。在AIM 2中，POL的角色？通过新设计的基于SV40的质粒系统，将分析通过人类细胞中不同类型的DNA病变促进复制。在要研究的DNA病变中，是由细胞氧化DNA损伤引起的8-氧气（8-oxog）和胸腺醇（TG）。环上的N2-（3-羟基丙基-2'-脱氧核苷[（R）-3HOPDG]和笨重的反式-4-羟基-2-hydroxy-2-non-enal--脱氧瓜烷（HNE-DG），这是由于n2-dg的反应和N2-dG的反应而产生的，并从n2-dg产生了admers oftical and radicer oftical and trad indicr and trad indicr and trad indicr ins free trids，多环苯甲酸[A] pyrene 7,8-二醇9,10-环氧化物（BPDE）N2-DG，导致对环境污染物和致癌物的暴露导致，我们会在AIM 3中对POL和Cryst-s and Cryst and cog and Crys and cog and Crys and cog and Crys， N2-DG的（r）-3HOPDG，HNE和BPDE的次数都可以通过其他DNA POL插入N2-DG加合的能力。从暴露于化学和环境致癌物中，POL？通过保持突变速率较低，对基因组稳定性产生重大影响，从而降低了人类癌变的发生率。拟议的研究与癌症生物学和病因高度相关，因为它们将揭示人类细胞如何最大程度地减少DNA病变的诱变和致癌潜力。

项目成果

Yeast 9-1-1 complex acts as a sliding clamp for DNA synthesis by DNA polymerase ε.

• DOI: 10.1016/j.jbc.2022.102727
• 发表时间: 2023-01
• 期刊: The Journal of biological chemistry
• 作者: Acharya N;Prakash L;Prakash S
• 通讯作者: Prakash S

Role of human DNA polymerase κ in extension opposite from a cis-syn thymine dimer.

人类 DNA 聚合酶 γ 在与顺式胸腺嘧啶二聚体相反的延伸中的作用。

• DOI: 10.1016/j.jmb.2011.02.042
• 发表时间: 2011
• 期刊: Journal of molecular biology
• 影响因子: 5.6
• 作者: Vasquez-DelCarpio,Rodrigo;Silverstein,TimothyD;Lone,Samer;Johnson,RobertE;Prakash,Louise;Prakash,Satya;Aggarwal,AneelK
• 通讯作者: Aggarwal,AneelK