Recognition of Environmental Carcinogen-DNA lesions by NER Proteins

NER 蛋白识别环境致癌物 DNA 损伤

• 批准号: 9057542
• 负责人: Nicholas E Geacintov
• 金额: $ 35.34万
• 依托单位: NEW YORK UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-08-01 至 2019-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9057542
• 关键词: Adenine; Affect; Affinity; Amaze; Aromatic Polycyclic Hydrocarbons; Asthma; Base Sequence; Benzo(a)pyrene; Binding; Biochemical; Biological Markers; Cancer Etiology; Carcinogens; Cell Extracts; Characteristics; Chemical Structure; Chicago; Cisplatin; Collaborations; Complex; Computational Technique; DNA; DNA Adducts; DNA Binding; DNA Damage; DNA Maintenance; DNA Repair; DNA lesion; Disease; Environment; Environmental Carcinogens; Environmental Pollution; Epoxy Compounds; Etiology; Excision; Exhibits; Exposure to; Food; Fossil Fuels; GTF2H1 gene; Glycols; Guanine; Health; Human; Illinois; Individual; Laboratories; Lesion; Libraries; Lung; Lung diseases; Malignant Neoplasms; Malignant neoplasm of lung; Methodology; Methods; Molecular Conformation; Molecular Models; Nucleotide Excision Repair; Nucleotides; Oxidation-Reduction; Oxidoreductase; Particulate; Process; Property; Proteins; Quinones; Reactive Oxygen Species; Resistance; Risk; Roentgen Rays; Shapes; Site; Smoker; Specificity; Structure; Structure-Activity Relationship; Surface Plasmon Resonance; Surgical incisions; System; Techniques; Testing; Thermodynamics; Tissues; Tobacco smoke; Water; Workplace; adduct; base; crosslink; exposed human population; gel electrophoresis; helicase; human DNA; human disease; improved; insight; molecular modeling; prototype; repaired; research study; superfund site; transcription factor TFIIH; urban area; wasting

DESCRIPTION (provided by applicant): The combustion of fossil fuels generates polycyclic aromatic hydrocarbons (PAH) that are ubiquitous and potentially cancer-causing environmental contaminants. PAH are found at toxic waste dumps and superfund sites, in airborne particulates, in our food and water, as well as in tobacco smoke. PAH compounds are believed to contribute to the higher rates of lung and other cancers that have been documented in residents of polluted urban areas and smokers. The PAH are biologically inactive but are metabolically activated to reactive diol epoxides that covalently bind to guanine and adenine in DNA to form stable, pre- mutagenic DNA adducts. Such forms of DNA damage accumulate in tissues of people exposed to PAH- contaminated environments, who are thus at risk of developing respiratory diseases and cancer. Not all DNA adducts are equally threatening to human health. Many of them can be removed by the human DNA repair mechanism called nucleotide excision repair (NER). However, the activity of the NER system is variable: some DNA lesions are slowly repaired, while some others are resistant to NER. The accumulation of such persistent DNA damage enhances the risk of developing diseases of the lung such as asthma, lung cancer, and other disorders. The exact structural features that render certain PAH-DNA adducts NER-resistant, are poorly understood. Among the first mammalian NER factors that recognize and bind to NER substrates is the heterodimeric XPC-RAD23B protein. The identification of the lesions occurs in a two-step (bipartite manner): (1) recognition by XPC-RAD23B, and (2) a subsequent verification step that involves the helicase activity of XPD in TFIIH, a multi-protein NER factor that binds to the XPC-DNA complex. The feasibility of this project is based on a previously developed, extensive library of different PAH-DNA adducts that exhibit the full spectrum of NER activities, from fully resistant to fully susceptible. The objecties are to elucidate the structural features of DNA lesions that abrogate or promote their recognition and repair. Aim 1 is focused on developing surface plasmon resonance and other methods (footprinting, gel electrophoresis) for studying XPC and TFIIH protein-DNA interactions utilizing a set of well characterized benzo[a]pyrene - diol epoxide guanine lesions (BP-G) in two different base sequence contexts. In one of these, the BP-G lesions are either moderate-to-good NER substrates; in the other sequence, a single nucleotide opposite the BP-G, lesion is missing, and the same BP-G duplexes are fully resistant to NER in human cell extracts. In Aim 2, these methodologies will be applied to investigate the mechanisms of the initial XPC- RAD23B and TFIIH-DNA binding phenomena utilizing different bulky and small NER-proficient and NER- resistant PAH-guanine and -adenine DNA adducts. Mechanistic insights will be gained by exploring the local thermodynamic destabilization of DNA caused by the lesions using NMR methods, and by molecular modeling and computational techniques.

描述（由申请人提供）：化石燃料的燃烧会产生多环芳烃（PAH），它们无处不在且潜在的引起癌症的环境污染物。在有毒的废物垃圾场和超级基金地点，空气中的颗粒物，我们的食物和水以及烟草烟雾中发现了PAH。据信，PAH化合物有助于较高的肺部和其他癌症，这些肺部已记录在受污染的城市地区和吸烟者的居民中。 PAH在生物学上是非活性的，但被代谢激活为反应性二醇环氧化物，它们共价结合DNA中的鸟嘌呤和腺嘌呤，形成稳定的诱变前DNA加合物。这种形式的DNA损伤积聚在暴露于PAH被污染环境的人的组织中，因此，他们有呼吸道疾病和癌症的风险。 并非所有DNA加合物都同样威胁到人类健康。其中许多可以通过称为核苷酸切除修复（NER）的人类DNA修复机制去除。但是，NER系统的活性是可变的：一些DNA病变缓慢修复，而另一些DNA病变对NER具有抗性。这种持续性DNA损伤的积累增加了肺部，肺癌和其他疾病等肺部疾病的风险。使某些PAH-DNA加合物NER NER的确切结构特征知之甚少。在识别和结合NER底物的第一个哺乳动物NER因子中，是异二聚体XPC-RAD23B蛋白。病变的识别以两步（双分化方式）发生：（1）通过XPC-RAD23B识别，（2）随后的验证步骤，涉及XPD在TFIIH中的解旋酶活性，TFIIH，一种与XPC-DNA复合物结合的多蛋白质因子。该项目的可行性是基于先前开发的，广泛的不同PAH-DNA加合物的库，这些图书馆表现出各种NER活动，从完全抵抗力到完全易感。这些物体是要阐明消除或促进其识别和修复的DNA病变的结构特征。 AIM 1的重点是开发表面等离子体共振和其他方法（占地面积，凝胶电泳），用于研究XPC和TFIIH蛋白-DNA相互作用，利用一组表征良好的苯并[a] pyrene-Diol Epodoxide瓜烷内鸟嘌呤鸟嘌呤鸟嘌呤病变（BP-G）在两种不同的基本序列中。在其中之一中，BP-G病变要么是中度到良好的NER底物。在另一个序列中，在BP-G的对面，缺少病变，并且相同的BP-G双链体对人类细胞提取物中的NER完全抗性。在AIM 2中，这些方法将用于研究最初的XPC-RAD23B和TFIIH-DNA结合现象的机制，利用不同的笨重和小型NER且耐Ner的PAH-鸟嘌呤和腺苷DNA加合物的机制。通过探索使用NMR方法以及分子建模和计算技术引起的DNA的局部热力学不稳定，将获得机械洞察力。