ROLE OF RNA POLYMERASE IN BACTERIAL DIFFERENTIATION

RNA 聚合酶在细菌分化中的作用

• 批准号: 3269318
• 负责人: Richard Marc Losick
• 金额: $ 32.77万
• 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/3269318
• 关键词: Bacillus subtilis; DNA directed RNA polymerase; Escherichia coli; bacterial genetics; cell differentiation; endonuclease; gene expression; gene mutation; genetic library; genetic manipulation; genetic mapping; genetic regulation; genetic transcription; nucleic acid sequence; regulatory gene; ribosomes; spectrometry

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 在孢子形成过程中，基因表达受到调控。 革兰氏阳性细菌枯草杆菌。具体来说，我们建议 操纵基因的表达，这些基因的转录是在 开始产孢子或处于发育的中间阶段，通过 启动子突变、调控突变和基因外突变的分离 抑制子突变。这些突变的影响将在 通过将基因和操纵子融合到大肠杆菌的lacZ基因和 用纯化形式的大肠杆菌进行体外转录研究 在体外，通过对纯化形式的B。 枯草杆菌RNA聚合酶全酶。这个项目的一个主要目的是 了解Spoo基因产物在Spoo基因产物启动阶段的作用 产孢子，这一问题将通过对 两个基因的启动子，它们的转录在开始时被诱导 孢子形成，并通过对新的RNA聚合酶信号因子的研究 这决定了他们的认可度。我们还将研究对 第三个基因，其诱导与遗传和形态事件相耦合 发生在产孢子的中间阶段。此外，我们还将 开发转座子Tn917的融合生成衍生物，将 枯草芽孢杆菌基因与lacZ基因的体内转录融合 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子基因。最后，我们 建议对RNA聚合酶西格玛因子模型进行遗传学检验 识别保守核苷酸的序列特异性DNA结合蛋白 同源启动子“-35”和“-10”区域的序列信号 通过O-37菌种特异性突变株的分离 枯草杆菌西格玛因子。预计这些研究将 高可及原核细胞模型系统中的细胞分化 可提供对正常和异常问题的重要见解 在高等细胞中的分化。