Regulation of Bacterial Two-Component Signaling by Small Membrane Proteins

描述（由申请人提供）：双组分系统是细菌用于感知和响应其环境的主要信号转导模式之一。这些回路在使细菌适应不同的生长条件、控制发育程序和启动致病生活方式方面发挥着关键作用。典型的双组分系统由两种蛋白质组成，上游的组氨酸激酶通常参与信号检测，下游的响应调节器控制电路输出。 信息流通过将磷酰基从组氨酸激酶转移到反应调节剂而发生。然而，双组分系统可以偏离这种简单的结构，并且具有额外的蛋白质组分或更复杂的磷酸转移途径。在以前的工作中，我们发现一个小的47个氨基酸的膜蛋白，MgrB，抑制PhoQ的活动。由于MgrB的表达是由PhoP激活的，因此该蛋白质作为PhoQ/PhoP回路中负反馈回路的一部分发挥作用。PhoQ还受到另一种小膜蛋白SafA的刺激，其表达由酸响应性EvgS/EvgA双组分系统控制。MgrB和SafA是最近发现的一组调节组氨酸激酶功能的小疏水蛋白。在这项提案中，我们将结合联合收割机荧光显微镜跟踪电路行为在单细胞中，与遗传分析和建模，以探讨MgrB和SafA的作用，在PhoQ/PhoP信号。我们将确定这些蛋白质对镁、抗菌肽和pH刺激下PhoQ/PhoP信号转导动力学和输入输出行为的影响，比较天然E.大肠杆菌分离株，表征新的输入信号，通过MgrB调制PhoQ/PhoP电路，并研究与MgrB-PhoQ-PhoP通路失调相关的生长缺陷。

DESCRIPTION (provided by applicant): Two-component systems are one of the primary modes of signal transduction that bacteria use to sense and respond to their environment. These circuits play a key role in enabling bacteria to adapt to diverse growth conditions, control developmental programs, and initiate pathogenic lifestyles. The prototypical two-component system consists of two-proteins, an upstream histidine kinase that is usually involved in signal detection, and a downstream response regulator, which controls the circuit output. Information flow occurs by transfer of a phosphoryl group from the histidine kinase to the response regulator. Two-component systems can deviate from this simple architecture, however, and have additional protein components or more complex phosphotransfer paths. In previous work we discovered that a small 47 amino acid membrane protein, MgrB, inhibits PhoQ activity. Since MgrB expression is activated by PhoP, the protein functions as part of a negative feedback loop in the PhoQ/PhoP circuit. PhoQ is also stimulated by SafA, another small membrane protein, whose expression is controlled by the acid-responsive EvgS/EvgA two component systems. MgrB and SafA are among a group of recently discovered small hydrophobic proteins that modulate histidine kinase function. In this proposal we will combine fluorescence microscopy to follow circuit behavior in single cells, with genetic analysis and modeling to explore the role of MgrB and SafA in PhoQ/PhoP signaling. We will determine the effect of these proteins on the dynamics and input-output behavior of PhoQ/PhoP signaling for stimulation with magnesium, antimicrobial peptides and pH, compare the behavior of the low pH response among natural E. coli isolates, characterize new input signals that modulate the PhoQ/PhoP circuit through MgrB, and study the growth defect associated with dysregulation of the MgrB-PhoQ-PhoP pathway.