AUTOREGULATION AND GLOBAL CONTROL OF THE CRP OPERON

CRP 操纵子的自动调节和全局控制

• 批准号: 3290108
• 负责人: MARTIN FREUNDLICH
• 金额: $ 9.8万
• 依托单位: STATE UNIVERSITY NEW YORK STONY BROOK
• 依托单位国家: 美国
• 财政年份: 1986
• 资助国家: 美国
• 起止时间: 1986-04-01 至 1989-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3290108
• 关键词: DNA binding protein; Escherichia coli; bacterial genetics; binding proteins; cell differentiation; cell growth regulation; cyclic AMP; gel electrophoresis; gene expression; genetic promoter element; genetic transcription; messenger RNA; molecular cloning; nucleic acid sequence; oligonucleotides; operon; point mutation; radioassay; site directed mutagenesis

Cyclic AMP is a key compound in many organisms involved in regulating numerous aspects of normal and abnormal cellular growth and differentiation. In most systems this control is mediated by a cyclic AMP binding protein. This protein is necessary for the biological function of cyclic AMP and, in some cases, the protein is also involved in regulating cyclic AMP synthesis. A knowledge of how the levels of the cyclic AMP binding protein is controlled is an important aspect of understanding how this key control element functions in the cell. We have begun a study of the regulation of the cyclic AMP binding protein (CRP) in E. coli. Our initial investigations suggest a unique mechanism for control of expression of the crp operon. This mechanism involves the activation by cyclic AMP and CRP of a divergently transcribed RNA that initiates close to the start of crp transcription but on the opposite strand. This RNA blocks the synthesis of crp mRNA. We wish to continue these studies using in vitro and in vivo techniques to prove that this novel mechanism regulates crp expression and to understand the molecular basis for its action. In vitro work will include site directed mutagenesis, the effect of the divergent RNA on transcription and experiments to test if the divergent RNA forms a duplex with the 5' end of crp mRNA. In vivo experiments will focus on the use of cloned fragments of the crp and divergent promoter to assess the role of the divergent transcription in crp regulation in vivo.

环磷酸腺苷是许多生物体中参与调节 正常和异常细胞生长的许多方面， 分化 在大多数系统中，这种控制是由环AMP介导的。 结合蛋白 这种蛋白质是必需的生物功能， 环AMP，在某些情况下，该蛋白质也参与调节 环AMP合成。 了解环磷酸腺苷水平如何 结合蛋白是如何被控制的， 该键控元件在单元中起作用。 我们已经开始研究 环磷酸腺苷结合蛋白（CRP）在E.杆菌 我们 最初的研究表明了一种控制表达的独特机制 crp操纵子。 这一机制涉及环AMP的激活 和CRP的发散转录的RNA，启动接近开始 但在相反的链上。 这种RNA阻断了 crp mRNA的合成。 我们希望继续这些研究， 和体内技术来证明这种新的机制调节CRP 表达和理解其作用的分子基础。 体外 工作将包括定点诱变，发散的影响， RNA对转录的影响以及测试趋异RNA是否形成 与crpmRNA的5'末端形成双链体。 体内实验将集中在 使用CRP和趋异启动子的克隆片段来评估 趋异转录在体内CRP调节中的作用。