Mechanism of CsrA-Mediated Global Control

描述(由申请人提供)：将通过研究全球调控中的一个新范式--大肠杆菌的碳储存调控(CSR)系统来寻求对转录后调控机制的洞察。CSR包括CSRA，一种RNA结合蛋白，调节靶mRNAs的翻译和/或稳定性。CsrB和CSRC是非编码调控RNA，通过隔离这种蛋白来对抗CSRA，而CsrD是一种特异性针对CsrB和CSRC的蛋白质，使其被RNase E降解。在大肠杆菌中，CSRA在广泛的范围内影响新陈代谢、生理、运动和多细胞行为，抑制从指数生长向稳定期转变过程中表达的某些基因，并激活指数阶段表达的各种基因。CSRA同源物广泛分布于真细菌中，调节植物和动物病原菌中毒力因子的表达。因此，拟议的研究还将提供对细菌生理调节和发病机制的基本了解，并可能为细菌感染提供新的治疗方法。这一建议的具体目的是：1)阐明CSRA激活或抑制基因表达的分子机制。这一目标将包括对CSRA介导的自动调节的分析，以及对影响CSRA靶标转录本稳定性的因素的调查。2)结合生物信息学、基因组学、分子遗传学和生物化学方法建立CSR全球调控子。3)CSRA-RNA络合物的化学计量和结构表征。这一目标将包括模型CSRA靶RNA的核磁共振结构测定，以及通过X射线结晶学确定CSRA-RNA络合物的结构。4)阐明CsrD作用的分子机制。我们将进行实验，以阐明CsrD专门针对CsrB和CSRC进行RNaseE降解的机制。这些研究的长期目标是全面了解CSR系统的调节成分、遗传电路、分子机制和生物功能。通过研究全球遗传调控中的一个新范式--大肠杆菌的碳储存调控(CSR)系统，将寻求对转录后调控机制的洞察。CSR在广泛的范围内控制细菌的代谢、生理、运动和生物膜的发育，并调节植物和动物病原体中毒力因子的表达。因此，拟议的研究将提供对细菌生理调节和发病机制的基本了解，并可能为细菌感染提供新的治疗方法。

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.