Mechanism of CsrA-Mediated Global Control

描述（由申请人提供）：通过研究大肠杆菌的碳储存调节（Csr）系统，将深入了解转录后调节机制。Csr包括CsrA，一种调节靶mRNA的翻译和/或调节靶mRNA的稳定性的RNA结合蛋白。CsrB和Csrc是非编码调节RNA，其通过隔离CsrA蛋白来拮抗该蛋白，CsrD是特异性靶向CsrB和Csrc以被RNase E降解的蛋白。在大肠在大肠杆菌中，CsrA广泛影响代谢、生理、运动和多细胞行为，抑制从指数生长期到稳定期生长过渡期间表达的某些基因，并激活在指数期期间表达的各种基因。CsrA同源物广泛分布于真细菌中，并调节植物和动物病原体中毒力因子的表达。因此，拟议的研究还将提供对细菌生理学和发病机制的调节的基本理解，并可能为细菌感染提出新的治疗方法。本研究的具体目的是：1）阐明CsA激活或抑制基因表达的分子机制。这一目标将包括CsRA介导的自动调节的分析，以及调查的因素，影响CsRA靶转录的稳定性。2)使用生物信息学、基因组学、分子遗传学和生物化学方法的组合建立Csr全局调节子。3)CsrA-RNA复合物的化学计量和结构表征。这一目标将包括一个模型CsrA靶RNA的NMR结构测定，以及CsrA-RNA复合物的结构，通过X射线晶体学。4)阐明CsRD作用的分子机制。我们将进行实验以阐明CsrD特异性靶向CsrB和Csrc以被RNase E降解的机制。这些研究的长期目标是充分了解Csr系统的调控成分、遗传电路、分子机制和生物学功能。深入了解转录后调控机制将寻求通过全球遗传调控，大肠杆菌的碳储存调控（Csr）系统的新范式的研究。Csr在植物和动物病原体中广泛控制细菌的代谢、生理、运动和生物膜的形成，并调节毒力因子的表达。因此，拟议的研究将提供细菌生理学和发病机制的调节的基本理解，并可能提出新的细菌感染的治疗方法。

DESCRIPTION (provided by applicant): Insight into post-transcriptional regulatory mechanisms will be sought through the study of a novel paradigm in global regulation, the carbon storage regulatory (Csr) system of Escherichia coli. Csr includes CsrA, an RNA binding protein that regulates translation and/or modulates the stability of target mRNAs. CsrB and CsrC are non-coding regulatory RNAs that antagonize CsrA by sequestering this protein and CsrD is a protein that specifically targets CsrB and CsrC for degradation by RNase E. In E. coli, CsrA affects metabolism, physiology, motility and multicellular behavior on a broad scale, repressing certain genes expressed during the transition from exponential to stationary phase growth and activating various genes expressed during exponential phase. CsrA homologues are widely distributed among eubacteria and regulate the expression of virulence factors in both plant and animal pathogens. Thus, the proposed studies will also provide fundamental understanding of the regulation of bacterial physiology and pathogenesis, and may suggest novel therapeutic approaches for bacterial infections. The specifc aims of this proposal are: 1) Elucidate the molecular mechanisms by which CsrA activates or inhibits gene expression. This aim will include an analysis of CsrA-mediated autoregulation, as well as an investigation into the factors that influence the stability of CsrA target transcripts. 2) Establish the Csr global regulon using a combination of bioinformatic, genomic, molecular genetic and biochemical approaches. 3) Stoichiometric and structural characterization of CsrA-RNA complexes. This aim will include NMR structural determination of a model CsrA target RNA, as well as the structure of CsrA-RNA complexes by X-ray crystallography. 4) Elucidate the molecular mechanism of CsrD action. We will conduct experiments to elucidate the mechanism by which CsrD specifically targets CsrB and CsrC for degradation by RNase E. The long-range objectives of these studies are to fully understand the regulatory components, genetic circuitry, molecular mechanisms, and biological functions of the Csr system. Insight into post-transcriptional regulatory mechanisms will be sought through the study of a novel paradigm in global genetic regulation, the carbon storage regulatory (Csr) system of Escherichia coli. Csr controls bacterial metabolism, physiology, motility and biofilm development on a broad scale, and regulates the expression of virulence factors in both plant and animal pathogens. Thus, the proposed studies will provide fundamental understanding of the regulation of bacterial physiology and pathogenesis, and may suggest novel therapeutic approaches for bacterial infections.