Investigation of gliding motility in Bacteroidetes

致病细菌利用其细胞表面或分泌到环境中的蛋白质分子侵入并定植宿主生物。需要特定的运输机制将这些蛋白质分子从细胞内的合成点运输到细胞外。因此，这些蛋白转运体是细菌性疾病的重要致病因子。9型分泌系统（T9SS）是最近发现的一种细菌蛋白输出系统，它对引起严重牙周病的牙龈卟啉单胞菌和相关牙齿病原体的发病机制至关重要。这些细菌也与类风湿关节炎和阿尔茨海默病的病因有关。T9SS的进化起源在于滑翔细菌在固体表面上快速移动的机制。在这些细菌中，原t9ss输出与固体表面结合的蛋白质，然后沿着位于外膜的螺旋轨道沿着细胞体移动这些“粘附素”。最近的研究表明，T9SS需要一个能量输入来从外膜转运蛋白中提取底物蛋白。T9SS和滑行动力组件之间的相似性表明，这是通过使用滑行轨道的缩减版本来完成的。我们将利用黏附素在滑动细菌轨道上的运动作为实验模型来阐明T9SS运输中这一关键步骤的机制。我们使用滑翔运动模式生物，强johnsoniae黄杆菌的基本原理是，需要在完整的细胞中研究轨道运动的机械过程。先前的研究表明，在主动滑动的细胞中，单分子成像方法可以跟踪荧光标记的粘附素沿着细胞体在滑动轨道上的运动。相比之下，P. gingivalis T9SS的切割滑动轨迹太小，无法对轨迹上的运动进行光学解析以进行研究。滑动轨迹的组织和机制将通过细胞内成像和轨迹组件的生化、结构和计算特征来探索。从这项工作中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机制的理解。

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.