SRF OPERON--REGULATION AND ROLE IN GENETIC COMPETENCE

SRF 操纵子——遗传能力的调节和作用

• 批准号: 2883006
• 负责人: PETER ZUBER
• 金额: $ 23.67万
• 依托单位: OREGON GRADUATE INSTITUTE SCIENCE & TECH
• 依托单位国家: 美国
• 财政年份: 1992
• 资助国家: 美国
• 起止时间: 1992-02-01 至 2000-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2883006
• 关键词: Bacillus; Escherichia coli; bacterial genetics; fusion gene; gene mutation; genetic regulation; genetic transcription; genetic translation; immunoelectron microscopy; molecular cloning; nucleic acid sequence; operon

Bacteria possess complex signal transduction networks that govern the cells' response to environmental changes. In Bacillus subtilis, such a network controls processes that are activated when a growth-limiting environment is encountered. Systems which sense conditions of high cell density and starvation converge to activate the expression of the srf operon, encoding the enzyme complex that catalyzes the biosynthesis of the lipopeptide antibiotic, surfactin, and encoding a regulator of genetic competence, ComS, which is required for the expression of genes that function in DNA import. Studies of srf regulation and function will enhance our understanding of how bacteria regulate the production of toxins and antibiotics, and the process of genetic transformation, which is one way bacteria acquire genes that confer drug resistance. Mutations that alter the nutrition-dependent regulation of srf expression and sporulation, called ggr (glucose-glutamine-dependent regulation), result in an elevated concentration of sigma-H, encoded by spoOH and required for the production of the pheromone Csf (competence stimulating factor) which activates srf transcription. The ggr mutations also confer a Crs (catabolite resistant sporulation) phenotype. The ggr loci will be isolated and characterized to determine their roles in cell density- and nutritional control of srf expression. The comS gene, located within and out-of-frame with the srfB gene of the srf operon, is subject to regulation that is separate from that which controls srf transcription initiation. This regulation requires sinR, which also functions in the cell's decision to undergo sporulation or competence development. The region(s) of srf that functions in sinR- dependent control will be identified. Other mutations that effect specifically comS expression will be isolated and characterized by nucleotide sequence determination, and by introducing the mutations into sinR mutants or cells that overexpress sinR. The intracellular location and the cell type distribution (competent or non-competent cell) of ComS will be determined using anti-ComS antibody. The structure of ComS and its possible role in controlling the activity of DegU (a response regulator-like protein that functions in the contor of degradative enzyme production and competence), ComK (the transcriptional activator of competence genes) or MBA and B (homologues of ATP-dependent proteases and heat shock proteins, and negative regulators of ComK) will be investigated by mutational analysis and protein-protein crosslinking experiments. The identification of genes that encode ComS- binding proteins will be isolated using the yeast two-hybrid system.

细菌具有控制的复杂信号转导网络 细胞对环境变化的反应。在枯草芽孢杆菌中，这样的 网络控制增长限制时被激活的过程 遇到环境。感知高细胞条件的系统 密度和饥饿收敛以激活SRF的表达 操纵子，编码催化生物合成的酶复合物 脂蛋白抗生素，表面素和编码遗传调节剂 能力，coms，这是表达基因所必需的 DNA导入的功能。 SRF调控和功能的研究将 增强我们对细菌如何调节产生的理解 毒素和抗生素，以及遗传转化的过程， 是细菌获得赋予耐药性的基因的一种方式。突变 这改变了SRF表达的营养依赖性调节和 孢子形成，称为GGR（葡萄糖 - 谷氨酰胺依赖性调节），结果 在Sigma-H的浓度较高，由Spooh编码，需要 信息素CSF的产生（能力刺激因子） 激活SRF转录。 GGR突变也赋予CRS （抗分解代谢物孢子形成）表型。 GGR基因座将 孤立和表征以确定它们在细胞密度和 SRF表达的营养控制。 coms基因，位于内部和 与SRF操纵子的SRFB基因的框架外， 与控制SRF转录的法规 引发。该法规需要SINR，这也起作用 Cell的决定经历孢子形成或能力发展。这 SRF的区域，在依赖罪行控制中起作用将是 确定。其他影响coms表达的突变将会 通过核苷酸序列测定和以下特征 将突变引入过表达的SINR突变体或细胞 辛尔。细胞内位置和细胞类型分布（胜任 COM的非竞争细胞）将使用抗coms确定 抗体。 COM的结构及其在控制 DEGU的活性（响应调节剂样蛋白在功能中起作用 降解酶产生和能力的扭曲），comk（ 能力基因的转录激活因子）或MBA和B（ 依赖ATP的蛋白酶和热激蛋白和负调节蛋白 将通过突变分析和蛋白质 - 蛋白质进行研究 交联实验。编码coms-的基因的识别 结合蛋白将使用酵母双杂交系统分离。