ROLE OF RNA POLYMERASE IN BACTERIAL DIFFERENTIATION

这些研究的总体目标是确定其机制 孢子形成过程中基因表达受到调控 革兰氏阳性细菌枯草芽孢杆菌。 具体来说，我们建议 操纵其转录被诱导的基因的表达 孢子形成的开始或发育的中间阶段 启动子突变、调节突变和基因外突变的分离 抑制突变。 这些突变的影响将在 通过使用基因和操纵子融合到大肠杆菌的 lacZ 基因和体内 通过纯化形式的大肠杆菌进行体外转录研究 并通过纯化形式的 B. 进行体外转录研究。 枯草芽孢杆菌RNA聚合酶全酶。 该项目的主要目标是 了解 spoO 基因产物在起始阶段的作用 孢子形成，这个问题将通过对 两个基因的启动子，其转录在启动时被诱导 孢子形成，并通过对新型 RNA 聚合酶信号因子的研究 决定了他们的认可。 我们还将审查以下方面的监管 第三个基因，其诱导与遗传和形态事件相结合 发生在孢子形成的中间阶段。 此外，我们将 开发转座子 Tn917 的融合生成衍生物 在体内创建枯草芽孢杆菌基因与 lacZ 基因的转录融合 大肠杆菌或费氏弧菌的 luxA 和 luxB 基因作为新工具 用于快速分析多种类型的孢子形成基因。 最后，我们 提议从基因角度测试RNA聚合酶西格玛因子的模型 识别保守核苷酸的序列特异性 DNA 结合蛋白 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 枯草芽孢杆菌西格玛因子。 预计这些研究将针对 高度可访问的原核模型系统中的细胞分化 可以为正常和异常的问题提供重要的见解 高等细胞的分化。

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.