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: 3290106
• 负责人: MARTIN FREUNDLICH
• 金额: $ 8.05万
• 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 1994-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3290106
• 关键词: DNA binding protein; Escherichia coli; bacterial genetics; binding proteins; cell differentiation; cell growth regulation; cyclic AMP; gene expression; genetic promoter element; genetic transcription; microorganism growth; molecular cloning; nucleic acid hybridization; nucleic acid sequence; operon; site directed mutagenesis

Cyclic AMP is a key compound in many organisms involved in regulation 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 low 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. Evidence suggests that autoregulation of crp in vivo also involves this mechanism. 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 of 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 transcript in crp regulation in vivo. Other systems will be examined to determine if this novel regulatory mechanism is involved in the control of the expression of additional genes.

环AMP是许多生物体中的一种关键化合物，参与了 调控细胞正常和异常生长的诸多方面 和差异化。在大多数系统中，这种控制是由 一种环状AMP结合蛋白。这种蛋白质是人体必需的 环AMP的生物学功能，在某些情况下，还包括蛋白质 也参与调节环状AMP的合成。一种知识 低水平的环磷酸腺苷结合蛋白被控制 是理解此键如何控制 元素在单元格中起作用。我们已经开始了一项关于 环磷酸腺苷结合蛋白在大肠杆菌中的调控。我们的 初步调查表明，有一种独特的机制可以控制 C反应蛋白操纵子的表达。这一机制涉及 环AMP和C反应蛋白对差异转录RNA的激活作用 它在接近CRP转录开始的时候启动，但在 相反的一股。这种RNA阻止了CRP mRNA的合成。 有证据表明，体内C反应蛋白的自动调节也包括 这个机制。我们希望通过体外实验继续这些研究。 和活体技术来证明这种新的机制调节 C反应蛋白的表达及其分子基础的研究 行动。在体外的工作将包括定点突变， 转录发散RNA的作用及实验研究 检测发散的RNA是否与CRP的5‘端形成双链 MRNA.体内实验将集中在克隆人的使用上 C反应蛋白和差异启动子片段的作用评估 体内C反应蛋白调控的差异转录本。其他系统 将被审查以确定这种新的监管机制 参与控制额外基因的表达。