Vibrio cholerae biofilms: structure, function, regulation and role in infection

描述（由申请人提供）：霍乱弧菌引起霍乱，是水生环境的自然居民。季节性霍乱暴发发生在疾病流行的地方，并可在世界范围内传播。霍乱弧菌形成生物膜的能力（即，基质封闭、表面相关的群落）对于其在流行病之间的水生生境中的生存至关重要，并且有利于流行病期间的宿主到宿主的传播。该提案的目的是提高我们对生物膜基质成分、生物膜形成的机制和调控以及它们在霍乱弧菌生物学中的重要性的理解。在目标1中，我们将重点了解生物膜基质蛋白RbmA，RbmC和Bap1的功能。我们将确定RbmA和VPS相互作用的分子基础，并确定RbmA蛋白水解的分子决定因素和后果。我们将测试RbmC和Bap1是否与VPS结合，并确定对其功能重要的区域。我们还将确定这些蛋白质如何对生物膜的机械性能做出贡献，以及通过超分辨率显微镜确定每个基质成分的位置。在目标2中，我们将确定由调节蛋白，VpsR和VpsT，在生物膜形成的机制。我们将确定传感器组氨酸激酶磷酸化VpsR，并确定生物膜发育过程中VpsR的磷酸化动力学。我们还将确定生物膜形成过程中VpsT和VpsR的直接靶点，并确定VpsR和VpsT调节子成员在霍乱弧菌传播和传播到环境中的贡献。阐明霍乱弧菌生物膜形成的机制将有助于未来控制霍乱弧菌的策略 霍乱流行病，并促进确定用于在感染期间对抗病原体的新型药物靶标。

DESCRIPTION (provided by applicant): Vibrio cholerae causes the disease cholera and is a natural inhabitant of aquatic environments. Seasonal cholera outbreaks occur where the disease is endemic and can spread worldwide. V. cholerae's ability to form biofilms (i.e., matrix-enclosed, surface-associated communities) is crucial for its survival in aquatic habitats between epidemics and is advantageous for host-to-host transmission during epidemics. The objective of this proposal is to improve our understanding of biofilm matrix components, the mechanisms and regulation of biofilm formation, and their importance in the biology of V. cholerae. In Aim 1, we will focus on understanding how biofilm matrix proteins RbmA, RbmC and Bap1 function. We will determine the molecular basis of RbmA and VPS interactions and determine molecular determinants and consequences of RbmA proteolysis. We will test whether RbmC and Bap1 bind to VPS and identify regions important for their function. We will also determine how these proteins contribute to the mechanical properties of biofilms, as well as the location of each matrix component by super resolution microscopy. In Aim 2, we will determine the mechanism used by the regulatory proteins, VpsR and VpsT, in biofilm formation. We will identify sensor histidine kinases that phosphorylate VpsR, and determine phosphorylation dynamics of VpsR during biofilm development. We will also identify direct targets of VpsT and VpsR during biofilm formation and ascertain the contribution of VpsR and VpsT regulon members in V. cholerae transmission and dissemination into the environment. Clarifying the mechanisms of biofilm formation in V. cholerae will prove useful for the development of future strategies for controlling cholera epidemics, and facilitate identification of novel drug targets for combating the pathogen during infection.