Biological Effects of DNA Adducts Formed by Nitroaromatic Compounds

硝基芳香族化合物形成的 DNA 加合物的生物学效应

基本信息

批准号：
8067870
负责人：
Ashis K Basu
金额：
$ 31.48万
依托单位：
UNIVERSITY OF CONNECTICUT STORRS
依托单位国家：
美国
项目类别：
财政年份：
1998
资助国家：
美国
起止时间：
1998-02-01 至 2015-01-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8067870
关键词：
1-Nitropyrene Affect Aphidicolin Base Pairing Biological Biological Assay Bypass Carcinogens Cell Extracts Cell Line Cells Complement Complex DNA DNA Adducts DNA Damage DNA-Directed DNA Polymerase Diesel Exhaust Embryo Environmental Pollutants Enzymes Escherichia coli Etiology Family Frequencies Gene Mutation Genes Genetic Goals Guanine Health Human In Vitro Investigation Kinetics Lesion Light Malignant Neoplasms Malignant neoplasm of lung Mammalian Cell Methods Mismatch Repair Modeling Molecular Mutagenesis Mutagens Mutation Nucleotides Oligonucleotides Oncogenes Pattern Plasmid Cloning Vector Play Polymerase Property Proteins Public Health Research Research Design Role Saccharomyces cerevisiae Sampling Single-Stranded DNA Site Sulfolobus solfataricus Techniques Testing Viral Vector Work adduct base ds-DNA genotoxicity in vivo inhibitor/antagonist insight interest kidney cell malignant breast neoplasm pollutant polymerization prototype public health relevance repaired

项目摘要

DESCRIPTION (provided by applicant): The goal of this application is to investigate the mechanism by which nitroaromatic carcinogens cause mutations. We shall focus on the major DNA adducts formed by the nitropyrenes, the primary mutagenic pollutants in diesel exhaust, but also plan to initiate work on the DNA adducts of 3-nitrobenzanthrone, a recently discovered potent mutagenic and carcinogenic contaminant in diesel. We hypothesize that a group of DNA adducts formed by these nitroarenes play important roles in the etiology of human cancer via the mutagenic effects in crucial sequences of cancer genes (e.g., p53). We also hypothesize that most of these mutations occur via error-prone bypass of the lesions by Y-family DNA polymerases. In order to test these hypotheses, we shall synthesize and characterize oligonucleotides containing the major guanine adducts of 1-nitropyene, 1,6- and 1,8-dinitropyere, and 3-nitrobenzanthrone in important gene sequences. These adduct-containing oligonucleotides will be used to construct single-stranded plasmid or viral vectors and mutagenicity and genotoxicity will be evaluated in Escherichia coli and mammalian cells. The effect of specific repair or replication proteins will be assessed by using repair-proficient and -deficient cell lines or inhibitors. The cellular studies will be complemented by investigating in vitro kinetics and fidelity of polymerization catalyzed by Dpo4 and four human Y- family DNA polymerases. Initially, we shall focus on the C8 guanine adduct of 1-nitropyrene. The translesion synthesis products will be sequenced to determine the mutagenic pattern caused by each DNA polymerase. Finally, duplex plasmid vectors with these adducts will be constructed, and progression of replication fork will be examined in human cell extracts to determine if each adduct is significant block to fork progression. Mutational types and frequencies in double-stranded DNA will be determined. Together, these studies will provide a deeper understanding of the mechanism of mutagenesis induced by the DNA adducts of these nitroaromatic carcinogens. PUBLIC HEALTH RELEVANCE: Nitroaromatic compounds have been detected in a variety of environmental samples, including diesel exhaust. These compounds are potent mutagens and carcinogens, and it has been suggested that some of these pollutants play a role in human lung and breast cancer. Therefore, investigation of the mechanism by which the nitroaromatic carcinogens cause mutations has high significance to public health.

描述（由申请人提供）：本申请的目的是研究硝基芳香族致癌物引起突变的机制。我们将重点关注硝基芘（柴油机尾气中的主要致突变污染物）形成的主要DNA加合物，同时还计划启动对3-硝基苯并蒽酮（最近在柴油中发现的强致突变和致癌污染物）的DNA加合物的研究。我们假设这些硝基芳烃形成的一组 DNA 加合物通过癌症基因关键序列（例如 p53）的诱变作用在人类癌症的病因学中发挥重要作用。我们还假设大多数突变是通过 Y 家族 DNA 聚合酶容易出错的绕过病变而发生的。为了检验这些假设，我们将合成并表征重要基因序列中含有 1-硝基芘、1,6-和 1,8-二硝基芘以及 3-硝基苯并蒽酮的主要鸟嘌呤加合物的寡核苷酸。这些含有加合物的寡核苷酸将用于构建单链质粒或病毒载体，并将在大肠杆菌和哺乳动物细胞中评估致突变性和遗传毒性。将通过使用修复良好和缺陷的细胞系或抑制剂来评估特定修复或复制蛋白的效果。细胞研究将通过研究 Dpo4 和四种人类 Y 家族 DNA 聚合酶催化的聚合的体外动力学和保真度来补充。首先，我们将重点关注 1-硝基芘的 C8 鸟嘌呤加合物。对跨损伤合成产物进行测序以确定每种 DNA 聚合酶引起的诱变模式。最后，将构建具有这些加合物的双链体质粒载体，并在人细胞提取物中检查复制叉的进展，以确定每个加合物是否显着阻碍复制叉的进展。双链 DNA 的突变类型和频率将被确定。总之，这些研究将更深入地了解这些硝基芳香族致癌物的 DNA 加合物诱导的诱变机制。公共健康相关性：在多种环境样品中检测到硝基芳香族化合物，包括柴油机尾气。这些化合物是强效诱变剂和致癌物，有人认为其中一些污染物在人类肺癌和乳腺癌中发挥作用。因此，研究硝基芳香族致癌物引起突变的机制对公众健康具有重要意义。