ROLE OF RNA POLYMERASE IN BACTERIAL DIFFERENTIATION

RNA 聚合酶在细菌分化中的作用

• 批准号: 3269320
• 负责人: Richard Marc Losick
• 金额: $ 34.81万
• 依托单位: HARVARD UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1976
• 资助国家: 美国
• 起止时间: 1976-02-01 至 1990-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3269320
• 关键词: Bacillus subtilis; DNA directed RNA polymerase; Escherichia coli; bacterial genetics; cell differentiation; developmental genetics; endonuclease; fusion gene; gene expression; gene mutation; genetic library; genetic manipulation; genetic mapping; genetic promoter element; genetic transcription; nucleic acid hybridization; nucleic acid sequence; regulatory gene; ribosomes; spectrometry; sporogenesis

The overall goal of these studies is to determine the mechanisms whereby gene expression is regulated during the process of spore formation in the Gram-positive bacterium Bacillus subtilis. Specifically, we propose to manipulate the exprerssion of genes whose transcription is induced at the onset of sporulation or at intermediate stages of development through the isolation of promoter mutations, regulatory mutations and extragenic suppressor mutations. The effects of these mutations will be analyzed in vivo by employing gene and operon fusions to the lacZ gene of E. coli and in vitro by means of transcription studies with purified forms of E. coli and in vitro by means of transcription studies with purified forms of B. subtilis RNA polymerase holoenzyme. A principal aim of this project is to understand the role of spoO gene products in the initiation phase of sporulation, a problem that will be addressed through studies on the promoters for two genes whose transcription is induced at the onset of sporulation, and through studies on the novel RNA polymerase signa factors that determine their recognition. We willalso examine the regulation of a third gene whose induction is coupled to genetic and morphological events occurring at an intermediate stage of sporulation. In addition, we will develop fusion-generating derivatives of the transposon Tn917 that will create transcription fusions in vivo of B. subtilis genes to the lacZ gene of E. coli or to the luxA and luxB genes of Vibrio fischeri as new tools for the rapid analysis of sporulation genes of many types. Finally, we propose to test genetically the model that RNA polymerase sigma factors are sequence-specific DNA binding proteins that recognize conserved nucleotide sequence signals in the "-35" and "-10" regions of their cognate promoters through the isolation of change-of-specificity mutants of the O-37 species of B. subtilis sigma factor. It is anticipated that these studies on cellular differentiation in a highly accessible procaryotic model system may provide significant insights into problems of normal and abnormal differentiation in higher cells.

这些研究的总体目标是确定 基因表达在孢子形成过程中受到调控， 革兰氏阳性菌枯草芽孢杆菌。 具体而言，我们建议 操纵基因的表达，这些基因的转录是在转录起始点诱导的。 孢子形成的开始或在中间阶段的发展，通过 启动子突变、调节突变和基因外突变的分离 抑制基因突变 这些突变的影响将在 通过基因和操纵子融合到E.杆菌和 通过纯化形式的E.杆菌 并通过用纯化形式的B的转录研究在体外进行。 枯草杆菌RNA聚合酶全酶。 该项目的一个主要目标是 了解spoO基因产物在启动阶段的作用， 孢子形成，这一问题将通过研究 两个基因的启动子，其转录在转录起始时被诱导。 孢子形成，并通过研究新的RNA聚合酶信号因子 这决定了他们的认可。 我们还将研究一个 第三基因，其诱导与遗传和形态事件偶联 发生在孢子形成的中间阶段。 此外，我们会 开发转座子Tn 917的融合衍生物， 在体内产生B的转录融合。将枯草杆菌基因转化为lacZ基因 大肠大肠杆菌或费氏弧菌的luxA和luxB基因作为新的工具 用于快速分析多种类型的产孢基因。 最后我们 建议从遗传学上测试RNA聚合酶σ因子是 识别保守核苷酸的序列特异性DNA结合蛋白 在其同源启动子的“-35”和“-10”区域中的序列信号 通过分离O-37物种的特异性改变突变体， 的B。枯草杆菌σ因子 预计这些研究对 在高度可及的原核模型系统中的细胞分化 可以提供对正常和异常的问题的重要见解 在更高的细胞中分化。