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DNA SEQUENCING OF UV INDUCED CHROMOSOMAL MUTATIONS

紫外线诱导的染色体突变的 DNA 测序

基本信息

批准号：
6635899
负责人：
Christopher W Lawrence
金额：
$ 26.32万
依托单位：
UNIVERSITY OF ROCHESTER
依托单位国家：
美国
项目类别：
财政年份：
1983
资助国家：
美国
起止时间：
1983-12-01 至 2004-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/6635899
关键词：
DNA damage DNA directed DNA polymerase DNA replication Escherichia coli bacterial genetics bacterial proteins carcinogen testing high performance liquid chromatography mutagen testing mutagens nuclear magnetic resonance spectroscopy nucleic acid sequence operon radiation carcinogen radiation genetics transfection /expression vector ultraviolet radiation

项目摘要

The long term goal of this work is primarily to understand the molecular mechanisms that produce mutations when mutagen-damaged DNA is replicated in E. coli, and secondarily to investigate other processes that promote tolerance of DNA damage. Although induced mutagenesis, and damage tolerance in general, is regulated very differently in E. coli and eukaryotes, basic enzymological processess are likely to be similar. Information of significance to human genetic diseases, such as cancer, can therefore be gained from studies employing the unrivalled experimental tools available with the bacterium. The aim of this project is to analyze the processes in vivo, by employing the analytical power of experiments in which vectors carrying defined and specifically located lesions are transfected into some of the many well characterized mutant strains that are available with the bacterium. Specific aims include: indentification of the gene products that are responsible for efficient replication past a T-T dimer in cells lacking DNA polymerase II, IV, and V and proofreading activity; identification of the MucB residues responsible for its differences in mutagenic properties comared to pol V; investigation of the RecA-independent recombination mechanism for the error free bypass of the T_T pyrimidine (6-4) pyrimidinone UV-photoproduct; measurement of the deamination rates in vivo of cytosine within a TC cis-syn cyclobutane dimer; and determination of the structure of duplex oligonucleotide molecules that contain a T_C pyrimidine (6-4) pyrimidinone adduct and the dewar isomers of the T-T and T-C (6-4) photoproducts, using nuclear magnetic resonance. The first of these aims is concerned with identifying the DNA polymerase or accessory factor capable of promoting replication past a T-T dimer in the absence of enzymes usually associated with SOS mutagenesis, whereas the second goal is to explore the structural reasons for the markedly different properties of related polymerases. The third aim is designed to explore what appears to be a novel, RecA independent, recombination process, while the last two investigate the reasons for the high mutability and specificity of two important UV photoproducts.

这项工作的长期目标主要是了解当诱变剂损伤的DNA在大肠杆菌中复制时产生突变的分子机制，其次是研究促进DNA损伤耐受性的其他过程。虽然在大肠杆菌和真核生物中，诱导诱变和损伤耐受性的调节方式非常不同，但基本的酶学过程可能是相似的。因此，对人类遗传疾病（如癌症）具有重要意义的信息，可以通过利用与细菌有关的无与伦比的实验工具进行研究而获得。该项目的目的是通过利用实验的分析能力来分析体内的过程，在实验中，将携带明确和特定位置病变的载体转染到细菌中可用的许多具有良好特征的突变菌株中。具体目标包括：鉴定在缺乏DNA聚合酶II、IV和V的细胞中负责通过T-T二聚体进行有效复制的基因产物和校对活性；与pol V相比，导致其致突变性差异的MucB残基的鉴定；不依赖reca的T_T嘧啶（6-4）嘧啶紫外光产物无误差旁路重组机理研究TC顺式-同步环丁烷二聚体胞嘧啶体内脱氨速率的测定；以及利用核磁共振测定含有T_C嘧啶（6-4）嘧啶酮加合物的双寡核苷酸分子的结构以及T-T和T-C（6-4）光产物的杜瓦异构体。第一个目标是确定在缺乏通常与SOS诱变相关的酶的情况下能够促进T-T二聚体复制的DNA聚合酶或辅助因子，而第二个目标是探索相关聚合酶显著不同特性的结构原因。第三个目的是探索一种新颖的，独立于RecA的重组过程，而最后两个目的是研究两种重要的紫外光产物的高易变性和特异性的原因。

项目成果

期刊论文数量（15）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

Ultraviolet light induces different spectra of lacI sequence changes in vegetative and conjugating cells of Escherichia coli.

紫外线诱导大肠杆菌营养细胞和接合细胞中不同光谱的 lacI 序列变化。

DOI：
10.1016/0022-2836(88)90197-0
发表时间：
1988
期刊：
Journal of molecular biology
影响因子：
5.6
作者：
LeClerc,JE;Christensen,JR;Tata,PV;Christensen,RB;Lawrence,CW
通讯作者：
Lawrence,CW

UmuC function is not essential for the production of all targeted lacI mutations induced by ultraviolet light.

UmuC 功能对于紫外线诱导的所有靶向 lacI 突变的产生并不是必需的。

DOI：
10.1016/0022-2836(88)90198-2
发表时间：
1988
期刊：
Journal of molecular biology
影响因子：
5.6
作者：
Christensen,JR;LeClerc,JE;Tata,PV;Christensen,RB;Lawrence,CW
通讯作者：
Lawrence,CW

Efficient translesion replication in the absence of Escherichia coli Umu proteins and 3'-5' exonuclease proofreading function.

在没有大肠杆菌 Umu 蛋白和 3-5 核酸外切酶校对功能的情况下进行有效的跨损伤复制。

DOI：
10.1073/pnas.95.26.15519
发表时间：
1998
期刊：
Proceedings of the National Academy of Sciences of the United States of America
影响因子：
11.1
作者：
Vandewiele,D;Borden,A;O'Grady,PI;Woodgate,R;Lawrence,CW
通讯作者：
Lawrence,CW

U-U and T-T cyclobutane dimers have different mutational properties.

U-U和T-T环丁烷二聚体具有不同的突变特性。