Mechanotransduction in bacteria: how mechanical forces trigger virulence in Pseudomonas aeruginosa

铜绿假单胞菌（PA）是一种细菌病原体，其感染包括人类在内的多种宿主动物。PA是一个重要的公共卫生问题，是医院获得性感染、烧伤伤口感染和囊性纤维化病理学的主要原因。PA对许多常用抗生素具有高耐受性，这使得更好地了解PA毒力调节的明确需求。从历史上看，该领域一直专注于PA对营养素或信号分子等化学线索的反应。然而，它最近被证明，PA也调节毒力在其环境中的机械线索，并响应机械力的能力可能是基础PA的能力，以针对广泛的主机。毒力因子的表达是由皮利通过Chp系统的收缩诱导的，Chp系统类似于鞭毛趋化系统。然而，关于来自回缩的机械力如何传递到该系统尚不清楚。通过本提案，我想填补这一空白，并确定这一机械转导途径的物理机制，即，PA如何感知机械力并将其转化为基因表达的变化。在第一步中，我将扩展现有的组合光学捕获和荧光显微镜到几个位置和力反馈模式。在第二步中，我将在一个小时内对单个皮利施加受控的光学力，同时使用荧光监测毒力因子作为收缩力的函数。在第三步中，我将使用PA缺失突变体，以确定是否电机驱动的菌毛内的收缩或张力触发毒力通过激活Chp system.The这些研究所获得的信息将大大促进我们的理解PA作为一种机会致病菌和PA如何传感器固体表面作为一种方式来识别主机。

Pseudomonas aeruginosa (PA) is a bacterial pathogen that infects a wide variety of host animals including humans. PA is a significant public health concern as a leading cause of hospital acquired infections, burn-wound infections, and cystic fibrosis pathology. PA exhibits high tolerance for many commonly-used antibiotics, raising a clear need for a better understanding of PA virulence regulation. Historically, the field has focused on PAs response to chemical cues such as nutrients or signaling molecules. However, it has recently been demonstrated that PA also regulates virulence in response to mechanical cues in its environment and that the ability to respond to mechanical forces may underlie PAs ability to target a broad range of hosts. The expression of virulence factors is induced by the retraction of pili through the Chp system which resembles the flagellar chemotaxis system. However, there is nothing known about how mechanical forces from retraction are transmitted to this system. With the present proposal, I would like to fill this void and determine the physical mechanism of this mechanotransduction pathway, i.e., how PA senses mechanical forces and translates them to a change in gene expression.In the first step, I will extend an existing combined optical-trapping and fluorescence microscope to several position- and force-feedback modalities. In the second step, I will apply controlled optical forces on single pili over the course of an hour, while monitoring virulence factors as a function of retraction force using fluorescence. In a third step, I will use PA deletion mutants to determine if motor driven retraction or tension within the pilus triggers virulence by activating the Chp system.The information gained by these studies will substantially further our understanding of PA as an opportunistic pathogen and of how PA senses solid surfaces as a way of identifying hosts.