Phenotypic heterogeneity generated by histidine kinases based signaling networks

越来越多的研究表明，表型异质性是细菌群体的共同特征。该项目侧重于基于组氨酸激酶的信号网络的比较分析，以揭示产生表型异质性的设计原则。在第一期资助期内，研究了基于多输入组氨酸激酶的哈维弧菌群体感应级联与表型异质性的关系。我们的研究结果清楚地表明，这种由三种平行双功能组氨酸激酶和一个srna调节开关组成的信号级联具有导致表型异质性的固有特性。具体来说，群体感应组氨酸激酶的激酶与磷酸酶活性之比（受三种自诱导剂的不同影响）和主调控因子LuxR的拷贝数对表型异质性的程度至关重要。最后，将哈维氏弧菌暴露于人工高浓度的所有自诱导剂中，导致其转变为同质群体，其产生的生物膜比异质群体少。在第二个资助期，我们将重点研究大肠杆菌中多输入组氨酸激酶营养传感网络。该网络由两种组氨酸激酶和两种转运蛋白组成。它感知几种细胞内和细胞外代谢物的水平，通过蛋白质-蛋白质相互作用传递这些信息并诱导表型异质性。这种营养感知网络也存在于肠杆菌科的其他成员中，包括许多病原体，例如克雷伯氏菌、克罗诺杆菌、柠檬酸杆菌。所有可用的数据都强烈表明，大肠杆菌群体的个体通过这个网络感知可用的碳源，并制定下注对冲策略，以适应和生存在营养过渡期间。这些两面下注的策略可能导致不同的细胞命运，如生长和分裂细胞、生物膜形成细胞或持久性细胞。我们将通过定量分析和数学建模来比较群体感应和营养感应网络与表型异质性的关系，并特别关注鲁棒性、信号转导的分子机制和sRNA开关的作用。最后，我们将分析营养可利用性和代谢对大肠杆菌个体细胞命运的影响。此外，我们将研究尿路致病性大肠杆菌宿主定殖过程中营养感知和表型异质性的生物学意义。我们的目标是通过特定的饮食来操纵这些大肠杆菌种群，使其变得同质，从而更容易产生免疫反应或抗生素治疗。

More and more studies reveal that phenotypic heterogeneity is a common feature in bacterial populations. This project focuses on the comparative analysis of histidine kinases based signaling networks to unravel design principles that generate phenotypic heterogeneity. The multi-input histidine kinases based quorum sensing cascade in Vibrio harveyi was studied in correlation with phenotypic heterogeneity in the first funding period. Our results clearly indicate that this signaling cascade with three parallel bifunctional histidine kinases and a sRNA-regulated switch has the inherent property to cause phenotypic heterogeneity. Specifically, the ratio between kinase to phosphatase activities of the quorum sensing histidine kinases, which is differentially influenced by the three autoinducers, and the copy number of the master regulator LuxR were found to be crucial for the degree of phenotypic heterogeneity. Finally, exposure of V. harveyi to an artificial high concentration of all autoinducers caused the switch into a homogeneous population, which produced less biofilm than a heterogeneous one. We will focus on the multi-input histidine kinases nutrient sensing network in Escherichia coli in the second funding period. This network is composed of two histidine kinases and two transporters. It senses the level of several intra- and extracellular metabolites, transduces this information by protein-protein interactions and induces phenotypic heterogeneity. This nutrient sensing network is also found in other members of the Enterobacteriaceae including many pathogens, e.g., Klebsiella, Cronobacter, Citrobacter. All available data strongly suggest that the individuals of the E. coli population sense the available carbon sources through this network and develop bet-hedging strategies to adapt and survive during nutrient transitions. These bet-hedging strategies could result in different cellular fates, such as growing and dividing cells, biofilm-forming cells, or persister cells. We will compare the quorum sensing and the nutrient sensing networks in correlation with phenotypic heterogeneity by quantitative analysis and mathematical modeling with a specific focus on robustness, the molecular mechanism of signal transduction and the role of sRNA switches. Finally, we will analyze the impact of nutrient availability and metabolism on individual cellular fates of E. coli. In addition, we will investigate the biological significance of nutrient sensing and phenotypic heterogeneity during host colonization of uropathogenic E. coli. We aim to manipulate these E. coli populations by specific diets to become homogeneously and therefore, easier accessible to the immune response or antibiotic treatments.