MECHANISM OF INDUCTION OF VIRAL AND CELLULAR FUNCTIONS

病毒和细胞功能的诱导机制

• 批准号: 3272360
• 负责人: DAVID W MOUNT
• 金额: $ 19.11万
• 依托单位: UNIVERSITY OF ARIZONA
• 依托单位国家: 美国
• 财政年份: 1978
• 资助国家: 美国
• 起止时间: 1978-01-01 至 1993-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3272360
• 关键词: DNA footprinting; DNA repair; Escherichia coli; bacterial genetics; cell growth regulation; gene expression; gene induction /repression; gene mutation; genetic manipulation; genetic mapping; genetic transcription; genetic translation; microorganism genetics; molecular cloning; mutant; peptidases; protein sequence; protein structure function; recombinase; regulatory gene; transfection; ultraviolet radiation

The long term goal of this project is to understand the molecular basis of induction of the E. coli SOS response and the resulting mutagenesis. First, we will identify additional genes in the SOS regulon by cloning their regulatory regions upstream from a galK reporter gene in low and high copy number plasmids. LexA binding sites will be located by footprinting and genetic experiments. Since recent experiments have suggested that LexA protein may recognize a second type of operator, the interaction of LexA protein with such operators will be studied using methods previously established for the recA operator. Second, with the coming availability of a crystal structure for RecA protein, we will attempt to define domains which specify RecA functions in the SOS response by using information from recA mutants. This project will involve production of a database of sequence and functional information of already existing mutants. To add to this database, we will measure the cleavage of LexA and UmuD proteins in vivo in uninduced and UV-induced recA80, recA432 and recA433 mutants and analyze the cleavage properties of the purified mutant RecA proteins. Third, we will continue analysis of the signalling mechanism which induces the SOS response by activating RecA protein. We will map fragments of chromosomal DNA which prevent induction of the SOS response when these fragments are present on high copy number plasmids. We will identify the functions of the genes which are present as regulators of the SOS induction pathway. Finally, the relationship between induction of the SOS regulon and enhanced excision of pyrimidine dimers from the transcribed DNA strand will be investigated.

该项目的长期目标是了解 诱导大肠杆菌 SOS 反应以及由此产生的诱变。 首先，我们将通过克隆鉴定 SOS 调节子中的其他基因 它们的调节区域位于低和高水平的 galK 报告基因上游 拷贝数质粒。 LexA 结合位点将通过足迹法定位 和基因实验。 由于最近的实验表明 LexA 蛋白质可能识别第二种类型的操纵子，即 LexA 的相互作用 将使用以前的方法研究具有此类运算符的蛋白质 为recA操作员建立。 其次，随着即将到来的可用性 为了研究 RecA 蛋白的晶体结构，我们将尝试定义结构域 它通过使用以下信息在 SOS 响应中指定 RecA 功能： recA 突变体。 该项目将涉及建立一个数据库 现有突变体的序列和功能信息。 添加 在此数据库中，我们将测量 LexA 和 UmuD 蛋白的切割 体内非诱导和紫外线诱导的recA80、recA432和recA433突变体中 分析纯化的突变 RecA 蛋白的切割特性。 第三，我们将继续分析诱导的信号机制。 通过激活 RecA 蛋白来做出 SOS 反应。 我们将绘制以下片段 染色体 DNA 阻止 SOS 反应的诱导，当这些 片段存在于高拷贝数质粒上。 我们将确定 作为 SOS 诱导调节因子的基因的功能 途径。 最后，SOS 调节子的诱导之间的关系 并增强从转录 DNA 链中切除嘧啶二聚体 将被调查。