Sensory Mechanisms and Local Signaling in c-di-GMP-mediated Signal Transduction in Escherichia coli

大肠杆菌 c-di-GMP 介导的信号转导的感觉机制和局部信号传导

• 批准号: 314334421
• 负责人: Professorin Dr. Regine Hengge
• 金额: --
• 依托单位: Arbeitsgruppe Mikrobiologie
• 依托单位国家: 德国
• 项目类别: Priority Programmes
• 财政年份: 2016
• 资助国家: 德国
• 起止时间: 2015-12-31 至 2020-12-31
• 项目状态: 已结题
• 来源: https://gepris.dfg.de/gepris/projekt/314334421?language=en
• 关键词: Sensory; Mechanisms; Local; Signaling; di

Bacterial biofilms colonize diverse surfaces and are highly resistant against antibiotics or host immune systems. In these biofilms, cells are embedded in a self-produced protective matrix of extracellular biopolymers. Major biofilm matrix components in E. coli are amyloid curli fibres and the exopolysaccharide cellulose whose synthesis is under control of the stationary phase sigma factor RpoS (¿S) and the second messenger c-di-GMP. The latter is produced by diguanylate cyclases (DGC, with GGDEF domains) and is degraded by specific phosphodiesterases (PDE, with EAL domains). 25 of the 29 GGDEF/EAL domain proteins of E. coli K-12 are enzymatically active, with 12 being DGCs and 13 being PDEs. Many of these differentially expressed DGCs and PDEs are membrane-associated and their activities are controlled by N-terminal sensory domains.Using biofilm matrix production as a c-di-GMP target that provides for convenient phenotypes and readouts for genetic, molecular biological and biochemical analyses, the proposed project aims at clarifying the molecular mechanisms of signal sensing and processing at the primary and secondary levels of c-di-GMP signaling in E. coli. This includes (i) sensing environmental and cellular signals via the N-terminal sensory input domains of a novel class of PDEs, and (ii) sensing c-di-GMP via novel and unorthodox c-di-GMP effector mechanisms. In particular, the following signal perception and c-di-GMP effector mechanisms will be studied:- Redox control of cytoplasmic enzyme activity via disulfide bond formation and proteolytic processing in the periplasmic loop domain of the six E. coli PDEs with CSS-EAL domain architecture- Molecular structure, function and targets of YbjK, a DeoR-like transcription factor that seems to bind c-di-GMP as a G-quadruplex- Regulation of transcription by PdeL, a locally acting c-di-GMP-binding and ¿degrading 'trigger PDE' and transcription factor with a LuxR-EAL domain architecture- Structure and function of a putative c-di-GMP riboswitch in the intergenic region of the mRNA of the orf0317-pdeL operon, which down-regulates PdeL expression.Furthermore, the integration of these molecular processes within the large regulatory network that controls matrix production and thereby generates functionality and architecture of a biofilm will be elucidated.With its focus on sensory input in the control of DGCs and PDEs, c-di-GMP-sensing effector components and local c-di-GMP signaling, this project is expected to make a major contribution to achieving the goals of SPP 1879. Moreover, its results will have practical applications, e.g. in the development of novel anti-biofilm strategies or in synthetic biology.

细菌生物膜分布在不同的表面，对抗生素或宿主免疫系统具有高度的耐药性。在这些生物膜中，细胞被嵌入到细胞外生物聚合物的自我产生的保护性基质中。大肠杆菌的主要生物膜基质成分是淀粉样卷曲纤维和胞外多糖纤维素，其合成受固定相σ因子rpos(S)和第二信使c-di-GMP的控制。后者由二鸟苷环化酶(DGC，带GGDEF结构域)产生，并被特定的磷酸二酯酶(PDE，带EAL结构域)降解。在大肠杆菌K-12的29个GGDEF/EAL结构域蛋白中，有25个蛋白具有酶活性，其中12个是DGC，13个是PDE。许多差异表达的DGC和PDE是膜相关的，它们的活性受N-末端感受域的控制。利用生物膜基质的产生作为c-di-GMP的靶标，为遗传、分子生物学和生化分析提供方便的表型和读数，该项目旨在阐明在大肠杆菌中c-di-GMP信号的初级和次级水平的信号传感和处理的分子机制。这包括(I)通过一类新型PDE的N端感觉输入域来感知环境和细胞信号，以及(Ii)通过新的和非常规的c-di-GMP效应器机制来感知c-di-GMP。特别是，将研究以下信号感知和c-di-GMP效应机制：-通过二硫键的形成和具有css-EAL结构域结构的六个大肠杆菌PDE的周质环区的蛋白分解处理来氧化还原控制细胞质酶的活性-YbjK的分子结构、功能和靶点-YbjK是一种Deor样转录因子，似乎以G-四链的形式与c-di-GMP结合-PdeL转录的调节，一种局部作用的c-di-GMP结合和降解具有LuxR-EAL结构域的‘触发PDE’和转录因子--在orf0317-pdeL操纵子的基因间隔区的一个可能的c-di-GMP核糖开关的结构和功能，它下调PdeL的表达。此外，这些分子过程在控制基质产生从而产生生物膜的功能和结构的大的调控网络中的整合将被阐明。它的重点是控制DGC和PDE的感觉输入，c-di-GMP敏感的效应成分和局部的c-di-GMP信号，预计该项目将为实现战略计划1879的目标作出重大贡献。此外，它的结果将具有实际应用，例如在开发新的抗生物膜策略或合成生物学方面。