Mechanism of Caulobacter adhesion

描述(由申请人提供)：细菌对表面的附着在疾病中起着重要的作用，为表面的生物污染和生物膜的形成提供了关键的第一步。这项研究的总体目标是详细了解细菌黏附的机制，从黏附的生物物理到参与这一过程的细胞表面结构的生物合成的协调。这个项目利用了新月杆菌，在细胞的同一极以有序的方式合成黏附结构，使黏附的研究比大多数细菌更容易。黏附的初始阶段涉及鞭毛的运动和菌毛，黏附是通过合成一种Holdfast多糖来粘合的。病原菌也使用这些黏附结构，但它们在黏附中的作用机制尚不清楚。此外，单个Caulbacter细胞的粘附力是迄今为止测量到的最强的生物粘附力。这个项目有三个主要目标。第一个目标是将高度同步的培养与原子力显微镜、荧光显微镜和生物物理模型结合起来，以发展对粘连各个阶段的详细了解。特别是，这个目标将调查一种新发现的依赖表面接触的粘附性多糖输出的触发因素；这一机制也可能被病原体使用。第二个目的是确定Holdfast多糖的合成和附着蛋白的功能。描述了确定这些蛋白质的生化功能及其对粘附性的贡献的实验。第三个目的是阐明控制Holdfast合成的时机和极性定位的机制。在细胞周期中组成性表达Holdfast合成蛋白对Holdfast合成和黏附的时间的影响将被确定。将研究Holdfast合成和附着蛋白的定位及其相互依赖的定位，并确定它们定位所需的因素。细菌的黏附、多糖的生物合成、蛋白质和毒力因子的亚细胞定位是细菌致病的重要组成部分。从这个简单的模型系统的研究中获得的洞察力将适用于更复杂的细菌病原体，并将增强我们抑制它们的能力。

DESCRIPTION (provided by applicant): The adhesion of bacteria to surfaces plays an important role in disease, providing the critical first step in the biofouling of a surface and in biofilm formation. The general goal of this research is to reach a detailed understanding of the mechanisms of bacterial adhesion, from the biophysics of adhesion to the coordination of the biosynthesis of cell surface structures that participate in this process. This project takes advantage of the bacterium Caulobacter crescentus, in which adhesive structures are synthesized in an ordered fashion at the same pole of the cell, making the study of adhesion more amenable than in most bacteria. Initial stages of adhesion involve flagellar motility and pili, and adhesion is cemented by synthesis of a polysaccharide holdfast. Pathogenic bacteria also use these adhesive structures, but their mechanism of action in adhesion is poorly understood. In addition, the adhesive force of individual Caulobacter cells is the strongest ever measured for a biological adhesive. This project has three major aims. The first aim will use highly synchronized cultures coupled to atomic force microscopy, fluorescence microscopy, and biophysical modeling to develop a detailed understanding of the various stages of adhesion. In particular, this aim will investigate a newly discovered surface contact-dependent trigger of adhesive polysaccharide export; this mechanism may also be used by pathogens. The second aim is to determine the function of holdfast polysaccharide synthesis and attachment proteins. Experiments are described to determine the biochemical function of these proteins and their contribution to adhesion. The third aim is to elucidate the mechanisms that control the timing and polar localization of holdfast synthesis. The effect of constitutively expressing holdfast synthesis proteins during the cell cycle on the timing of holdfast synthesis and adhesion will be determined. The localization of holdfast synthesis and attachment proteins and their interdependence for localization will be studied, and factors required for their localization will be identified. Bacterial adhesion, polysaccharide biosynthesis, and subcellular localization of proteins and virulence factors are essential components of bacterial pathogenesis. Insight gained from the study of this simple model system will be applicable to more complex bacterial pathogens and will enhance our ability to inhibit them.