Cyclic di-GMP Regulation of PilB in Motility and Biofilm

是走还是留，这是细菌在任何环境中都必须做出的决定。要离开，它必须组装一个运动装置，并使附着在物体表面的结构失效。为了停留，它需要通过生产自己的建筑材料来在固体表面形成细菌生物膜，从而建立一个受保护的定居点。细菌的这一决定及其执行对我们的生活和社会产生了深远的影响，因为生物膜可以在许多关键的环境表面上形成。该研究项目在分子水平上探索细菌如何做出并执行这一决定，要么形成生物膜，要么通过使其运动性而移动。除了推进基础微生物学外，该项目将为研究生和本科生提供综合教与学的机会，以及跨学科的培训。此外，还将开展各种外展活动，以帮助我们的STEM劳动力多样化。本项目主要研究通用细菌第二信使环二gmp （cdG）对PilB的调控作用。PilB是一种atp酶，它为细菌IV型菌毛（T4P）的组装提供动力，作为在固体表面上运动的运动装置。此外，黄粘球菌T4P已被证明可以刺激胞外多糖（EPS）的产生，EPS是细菌生物膜的主要构建材料。因此，M. xanthus PilB （MxPilB）似乎同时促进了细菌的运动和生物膜的形成，这是两种相互替代和相互排斥的细菌生活方式。有证据表明，MxPilB和它的嗜热氯酸杆菌同源物（CtPilB）都通过一个保守的cdG结合结构域与cdG结合。研究人员提出了一个模型，其中cdG反向调节PilB的两个看似不相容和相反的功能。本项目的一部分使用黄原草在体内结合体外cdG结合试验对该模型进行测试。项目其余部分以CtPilB为模型蛋白，通过体外定量和结构分析，以及先进的成像平台，包括单粒子冷冻电镜和高速原子力显微镜，分析PilB与cdG的相互作用。这个项目的成功完成将提供对存在于许多生态系统和系统发育树的许多分支中的t4p运动细菌的生活方式决策的见解。该奖项反映了美国国家科学基金会的法定使命，并通过使用基金会的知识价值和更广泛的影响审查标准进行评估，被认为值得支持。

To go, or to stay, that is the decision a bacterium must make in any environment it encounters. To go, it must assemble a motility apparatus and disable structures for attachment to surfaces. To stay, it needs to establish a protected settlement by producing building materials of its own to form a bacterial biofilm on a solid surface. This decision and its execution by bacteria profoundly impact our lives and society because biofilms can form on so many critical environmental surfaces. This research project probes at the molecular level how a bacterium makes and executes this decision to either form biofilms or move by enabling its motility. Besides advancing basic microbiology, this project will allow integrated teaching and learning opportunities as well as interdisciplinary training of both graduate and undergraduate students. In addition, a variety of outreach activities will be performed to help diversify our STEM workforce.This project focuses on the regulation of PilB by the universal bacterial second messenger cyclic-di-GMP (cdG). PilB is an ATPase that powers the assembly of bacterial type IV pilus (T4P) as a motility apparatus for movement on solid surfaces. Additionally, Myxococcus xanthus T4P has been demonstrated to stimulate the production of exopolysaccharide (EPS) which is the major building materia of bacterial biofilms. M. xanthus PilB (MxPilB) would thus appear to promote both motility and biofilm formation, which are two alternate and mutually exclusive lifestyles of bacteria. Evidence indicates that MxPilB and its Chloracidobacterium thermophilum orthologue (CtPilB) both bind cdG using a conserved cdG-binding domain. The investigators propose a model wherein cdG inversely regulates the two seemingly incompatible and opposing functions of PilB. One part of this project uses M. xanthus to test this model in vivo combined with cdG binding assays in vitro. The rest of the project employs CtPilB as the model protein to analyze the interactions of PilB with cdG by quantitative and structural analysis in vitro as well as advanced imagining platforms including single-particle cryo-EM and high-speed atomic force microscopy. The successful completion of this project will provide insight into the lifestyle decision-making by T4P-motile bacteria that are present in numerous ecosystems and on many branches of the phylogenetic tree.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.