Finding the way: Sensory adaptation during bacterial mechanotransduction

摘要 感觉适应是一种短期的记忆过程，信号系统通过这个过程返回到它的 刺激前水平，尽管持续暴露在输入信号中。尽管很好地描述了 细菌趋化性，对适应如何在其他细菌信号中起作用知之甚少。 系统。更好地理解适应如何在这些系统中运行将提供新的 基础知识，并可确定新的治疗方法。我们专注于 机械传感是假单胞菌表面定植和感染的关键 铜绿假单胞菌(PA)是耐多药医院感染的主要原因，也是一种重要的 对健康的威胁。PA使用Pil-CHP机械传感系统来转换机械信号 推动抽搐运动和cAMP的产生来调节表面上的毒力程序 联系。适应如何在这个系统中发挥作用，或者说机械感知，还没有被探索。 我们建议剖析适应在PIL-CHP系统中的机制和作用，因为(I) 机械感觉适应系统的核心酶机制--甲基化 Pilk酶和去甲基化酶ChpB是保守的，但它的调节似乎是 有别于大肠杆菌趋化系统；(Ii)该系统提供了了解 如何部署适应以应对表面接触；和(3)PIL-CHP系统 对急性肺炎小鼠模型的毒力有贡献，表明其与 人类PA感染。我们发现Pilk是一种甲基酶，而ChpB则是一种 PilJ化学感受器的去甲基酶来控制这两种输出。与趋化性不同， 甲基酶和去甲基酶表现出相反的空间定位。我们的研究支持一个模型 其中Pilk甲基酶定位于滞后的极点，PilJ化学感受器将在那里 甲基化并准备被激活。相反，ChpB去甲基酶被招募到 通过与响应调节器PilG的相互作用领先杆位。需要PilG进行协调 TFP在前导极伸缩。这种本地化将导致暂时的和 在前导极有空间限制的PilJ去甲基化。PilJ的活动将会受到抑制， 潜在地促进了PilG重新定位到滞后的极点和反转。因此，感官 Pil-CHP系统中的适应与大肠杆菌趋化中的适应是根本不同的 因为它使用时间和空间线索。我们将对这一假设进行如下检验：目标1.链接 PilJ甲基化状态对PilJ活性和产量的影响。目标2.确定如何应对 调节子PilG调节ChpB去甲基酶。目的3.检测Pilk甲基酶是否 受c-di-GMP结合蛋白MapZ的调节，将抽动和鞭毛运动联系起来。

Abstract Sensory adaptation is a short-term memory process by which a signaling system returns to its pre-stimulus level despite ongoing exposure to the input signal. Although well-characterized in bacterial chemotaxis, little is known about how adaptation operates in the other bacterial signaling systems. A better understanding of how adaptation operates in these systems will provide new fundamental knowledge and could identify new therapeutic approaches. We focus on mechanosensing, which is critical for surface colonization and infection in Pseudomonas aeruginosa (PA), a leading cause of multi-drug resistant nosocomial infections and a significant health threat. PA uses the Pil-Chp mechanosensing system to transduce a mechanical signal that drives twitching motility and cAMP production to modulate a virulence program upon surface contact. How adaptation functions in this system, or mechanosensing in general, is unexplored. We propose to dissect the mechanism and role of adaptation in the Pil-Chp system because (i) The core enzymatic machinery of the mechanosensory adaptation system, the methylating enzyme PilK and demethylating enzyme ChpB, are conserved but its regulation appears to be distinct from the E. coli chemotaxis system; (ii) This system affords the opportunity to understand how adaptation is deployed in response to surface contact; and (iii) The Pil-Chp system contributes to virulence in a murine model of acute pneumonia, demonstrating its relevance to human PA infections. We discovered that PilK acts as a methylase and ChpB acts as a demethylase for the PilJ chemoreceptor to control the two outputs. Unlike in chemotaxis, the methylase and demethylase exhibit inverse spatial localization. Our studies support a model in which the PilK methylase localizes to the lagging pole, where the PilJ chemoreceptor would be methylated and poised to be activated. In contrast, the ChpB demethylase is recruited to the leading pole by interactions with the response regulator PilG. PilG is required for coordinating TFP extension and retraction at the leading pole. This localization would lead to temporally and spatially restricted PilJ demethylation at the leading pole. PilJ activity would be dampened, potentially facilitating PilG relocalization to the lagging pole and reversals. Thus, sensory adaptation in the Pil-Chp system is fundamentally different from adaptation in E. coli chemotaxis in that it uses temporal AND spatial cues. We will test this hypothesis as follows: Aim 1. Link PilJ methylation states to PilJ activity and outputs. Aim 2. Determine how the response regulator PilG regulates the ChpB demethylase. Aim 3. Test whether the PilK methylase is regulated by MapZ, a c-di-GMP binding protein, to link twitching and flagellar motility.