TEMPORAL AND SPATIAL GENE REGULATION--BACILLUS SUBTILIS

时空基因调控——枯草芽孢杆菌

• 批准号: 3468025
• 负责人: LEE R KROOS
• 金额: $ 9.1万
• 依托单位: MICHIGAN STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1989
• 资助国家: 美国
• 起止时间: 1989-12-01 至 1994-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3468025
• 关键词: Bacillus subtilis; DNA directed RNA polymerase; DNA footprinting; Escherichia coli; bacterial genetics; binding proteins; developmental genetics; fusion gene; gene expression; genetic promoter element; immunochemistry; microorganism growth; mutant; transcription factor

Sporulation of the bacterium Bacillus subtilis is a relatively simple developmental process that involves a highly ordered program of gene expression and morphological change. The goal of this research is to understand how gene expression is coupled to morphological change during Bacillus sporulation to produce the correct pattern of temporal and spatial gene expression. These studies are of broad significance since all organisms that undergo development must regulate gene expression system to uncover fundamental mechanisms involved in developmental gene regulation. Early in sporulation, the daughter chromosome are partitioned to two compartments, and the subsequent pattern of gene expression in each compartment is distinct. Genetic studies suggest that gene expression in the mother cell compartment is somehow coupled to events occurring in the forespore compartment. The properties of two recently discovered regulators of mother cell-specific gene expression provide tow plausible mechanisms for the coupling phenomenon. One of the regulatory proteins is a sigma subunit of RNa polymerase (called sigmak) that may be synthesized as an inactive precursor. Its processing to the active form could be coupled to forespore events. The other regulatory protein (called the 14kDa protein) is a non-sigma transcription factor whose inactivation has been proposed experiments are designed to test these hypotheses and to define the roles of these two proteins in temporal and spatial gene regulation. Specifically, 1) the binding site of the 14kDa protein in a promoter it represses will be mutated to see if the promoter now becomes active earlier and is uncoupled form forespore events, 2) antibody to the 14kDa protein will be prepared and used to determine its amount and location during sporulation, 3) the sink gene (encoding sigmak) will be fused to an inducible promoter and pro-sigmak will be tested for activity. Antibody to pro-sigmak will be prepared to test directly whether sigmak is made as a precursor during sporulation. If pre-sigmak is inactive, a truncated, active sigk gene will be fused to an induced promoter. The ability to produce pro-sigmak or sigmak at will in cells will permit testing of whether pro-sigmak processing is involved in the coupling between compartments.

芽胞形成枯草杆菌是一种相对简单的细菌 涉及高度有序的基因程序的发育过程 表达和形态变化。这项研究的目标是 了解基因表达是如何与形态变化相结合的 芽孢杆菌产孢量产生正确的时间和空间模式 基因表达。这些研究具有广泛的意义，因为 经历发育的生物必须调节基因表达系统，以 揭示涉及发育基因调控的基本机制。 在孢子形成的早期，子代染色体被分割成两个 以及随后每个隔间中基因表达的模式 车厢清晰可见。遗传学研究表明，人类的基因表达 母细胞隔间以某种方式与发生在 前孔隔间。最近发现的两个化合物的性质 母细胞特异性基因表达的调节提供了两种可能 耦合现象的机理。其中一种调节蛋白是 RNA聚合酶的一种sigma亚单位(称为sigmak)，可被合成 作为一种不活跃的前体。它对活动表单的处理可以是 再加上前瞻事件。另一种调节蛋白(称为 14 kDa蛋白)是一种非西格玛转录因子，其失活具有 已提出的实验旨在验证这些假设，并 明确这两种蛋白在时间和空间基因中的作用 监管。具体地说，1)14 kDa蛋白在 它抑制的启动子将发生突变，看看启动子现在是否成为 较早活跃并解偶联形成前孔事件，2)抗体 将制备14 kDa的蛋白质并用于测定其含量和 孢子形成过程中的定位，3)Sink基因(编码sigmak)将是 融合到一个可诱导的启动子和Pro-sigmak将进行活性测试。 将制备针对proSigmak的抗体来直接检测Sigmak是否为 在孢子形成过程中作为前驱物质制成。如果Pre-sigmak处于非活动状态，则 截短的活性SIGK基因将与诱导启动子融合。这个 能够在细胞中随意产生前Sigmak或Sigmak将允许 测试耦合中是否涉及Pro-sigmak处理 在车厢之间。