Characterization of non-canonical regulatory pathways in the Caulobacter NtrYX signaling system

项目摘要/摘要 所有细菌都必须感知并对新陈代谢和环境变化做出反应，才能在不同的 栖息地。这些自适应响应通常使用双分量信令系统来实现。两个- 组成信号系统由两个核心要素组成：感觉组氨酸激酶和反应 调整器。组氨酸激酶检测信号并使反应调节器磷酸化。总体而言, 反应调节子的磷酸化导致基因表达的变化。尽管典型的双组分 信令系统有些简单，但越来越明显的是，许多系统 复杂的感觉网络，结合了非典型特征和额外的调节组件。Ntry-NtrX (Ntryx)是一个复杂的感觉系统与许多⍺的不同表型相关的新兴例子- 变形杆菌。以前的研究已经在典型的两组分框架内研究了NtrYX 发信号。然而，最近的研究表明，在淡水和土壤细菌新月弯杆菌中， NtrYX系统更为精细，包含了一种未知的磷酸化来源和一个新的国家 监管机构，NtrZ。此外，遗传和转录数据表明，磷酸化和非磷酸化 NtrX具有独特的和生理上相关的活动。拟议的工作将调查综合监管 Crescentus NtrYX系统中多个水平的策略。这个项目的第一个目标将决定如何 NtrX的磷酸化影响其与DNA的结合和调节活性。为了更好地理解这一点 磷酸化受到控制，第二个目标将应用生化和结构方法来探索 国家和NtrZ调节器之间的相互作用。最后，第三个目标将探索另一条途径 调节磷酰化流动，采用候选驱动和无偏见的方法来识别体内来源 NtrX的磷酸化。总而言之，这些方法将把新月浑鱼建立为一个强大的模型 了解交叉调控通路如何控制双组分信号转导。我们的工作也将 阐明了不同⍺-变形杆菌中保守和重要的NtrYX系统，包括病原菌和 共生生物如流产布鲁氏菌和苜蓿中华根瘤菌。此外，该项目还将揭示新奇和 调整细菌中的信号转导途径的通用策略。

Project Summary/Abstract All bacteria must sense and respond to metabolic and environmental changes to thrive in diverse habitats. These adaptive responses are often achieved using two-component signaling systems. Two- component signaling systems are composed of two core elements: a sensory histidine kinase and response regulator. The histidine kinase detects a cue and phosphorylates the response regulator. In general, phosphorylation of response regulators leads to changes in gene expression. Although canonical two-component signaling systems are somewhat simple, it has become increasingly apparent that many systems are more complex sensory networks, incorporating atypical features and additional regulatory components. NtrY-NtrX (NtrYX) is an emerging example of a complex sensory system linked to diverse phenotypes in many ⍺- proteobacteria. Prior studies have investigated NtrYX within the framework of canonical two-component signaling. However, recent work reveals that, in the freshwater and soil bacterium Caulobacter crescentus, the NtrYX system is more elaborate, incorporating an unidentified source of phosphorylation and a novel NtrY regulator, NtrZ. In addition, genetic and transcriptional data suggest that phosphorylated and unphosphorylated NtrX have distinct and physiologically relevant activities. The proposed work will investigate integrated regulatory strategies in the C. crescentus NtrYX system at multiple levels. The first aim of this project will determine how NtrX phosphorylation affects its DNA-binding and regulatory activities. To better understand how this phosphorylation is controlled, the second aim will apply biochemical and structural approaches to probe the interaction between NtrY and the NtrZ regulator. Finally, the third aim will explore an additional pathway regulating phosphoryl-flow, employing candidate-driven and unbiased approaches to identify the in vivo source of NtrX phosphorylation. Together, these approaches will establish C. crescentus NtrYX as a powerful model for understanding how intersecting regulatory pathways can control two-component signaling. Our work will also shed light on conserved and important NtrYX systems in diverse ⍺-proteobacteria, including pathogenic and symbiotic organisms like Brucella abortus and Sinorhizobium meliloti. Moreover, this project will reveal novel and generalizable strategies for tuning signal transduction pathways in bacteria.