Investigation of gliding motility in Bacteroidetes

致病细菌利用细胞表面的蛋白质分子或分泌到环境中的蛋白质分子入侵并在宿主体内定居。需要特定的运输机制来将这些蛋白质分子从它们在细胞内的合成部位输送到细胞外部。因此，这些蛋白转运体是细菌疾病的重要致病因子。9型分泌系统(T9SS)是新近发现的一种细菌蛋白输出系统，在牙龈卟啉单胞菌和引起严重牙周病的相关牙科病原体的发病机制中起着至关重要的作用。这些细菌也与类风湿性关节炎和阿尔茨海默氏病的病因学有关。T9SS的进化起源在于滑动细菌在固体表面上快速移动所使用的机械。在这些细菌中，原T9SS输出与固体表面结合的蛋白质，然后在位于外膜的螺旋轨道上沿细胞体移动这些粘附素。最近的研究表明，T9SS需要能量输入才能从外膜转运蛋白中提取底物蛋白。T9SS和滑翔运动部件之间的相似性表明，这是使用滑行轨迹的简化版本来实现的。T9SS转运的这一关键步骤的机制将以滑行细菌中的粘附素在轨道上的运动为实验模型来阐明。我们使用滑动运动模式生物--强生黄杆菌--的基本原理是，需要在完整的细胞中研究轨道上运动的机械过程。已有研究表明，在主动滑动的细胞中，荧光标记的粘附素在滑动轨迹上沿细胞体的运动可以通过单分子成像方法来跟踪。相比之下，牙龈假单胞菌T9SS的滑行轨迹太小，无法在轨迹上移动，因此无法通过光学方法进行研究。滑行轨道的组织和机制将使用细胞内成像进行探索，并辅之以轨道组件的生化、结构和计算特征。这项工作获得的见解将用于指导实验直接测试T9SS功能。这些研究将大大增加我们对T9SS机制的了解。

Disease-causing bacteria invade and colonise the host organism using protein molecules on their cell surface or secreted into their environment. Specific transport mechanisms are needed to transport these protein molecules from their site of synthesis inside the cell to the cell exterior. These protein transporters are, thus, important pathogenicity factors in bacterial diseases. The Type 9 Secretion System (T9SS) is a recently-discovered bacterial protein export system that is essential for the pathogenesis of Porphyromonas gingivalis and related dental pathogens that cause severe periodontal disease. These bacteria have also been implicated in the etiology of rheumatoid arthritis and alzheimers disease. The evolutionary origin of the T9SS lies in the machinery used by gliding bacteria to move rapidly over solid surfaces. In these bacteria the proto-T9SS exports proteins that bind to the solid surface and then moves these `adhesins' along the cell body on helical tracks located in the outer membrane. It has recently been shown that the T9SS requires an energetic input to extract substrate proteins from the outer membrane transporter. Similarity between T9SS and gliding motility components suggests that this is accomplished using a cut-down version of the gliding track. The mechanism of this crucial step in T9SS transport will be elucidated using the movement of adhesins on tracks in gliding bacteria as our experimental model. Our rationale for using the gliding motility model organism, Flavobacterium johnsoniae, is that the mechanical process of movement on tracks needs to be studied in intact cells. It has previously been shown that the movements of fluorescently-labelled adhesins along the cell body on gliding tracks can be followed by single molecule imaging methods in actively gliding cells. By contrast, the cut-down gliding tracks of the P. gingivalis T9SS are too small for movement on the tracks to be optically resolved for study. The organisation and mechanism of gliding tracks will be probed using in-cell imaging complemented by biochemical, structural, and computational characterisation of track components. The insights obtained from this work will then be used to guide experiments to directly test T9SS function. These studies will substantially increase our understanding of the mechanism of the T9SS.