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

描述(申请人提供)：霍乱弧菌导致霍乱，是水生环境中的自然居民。季节性霍乱暴发发生在霍乱地方病流行的地方，并可在全球范围内传播。霍乱弧菌形成生物膜(即基质封闭的表面相关群落)的能力对其在疫情之间的水生生境中的生存至关重要，并有利于在疫情期间宿主至宿主的传播。这项建议的目的是提高我们对霍乱弧菌生物膜基质成分、生物膜形成机制和调控及其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蛋白降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影响生物膜的机械性能，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，我们将确定调节蛋白VpsR和VpsT在生物膜形成中使用的机制。我们将鉴定使VpsR磷酸化的传感器组氨酸激酶，并确定VpsR在生物膜发育过程中的磷酸化动力学。我们还将确定VpsT和VpsR在生物被膜形成过程中的直接靶点，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作用。阐明霍乱弧菌生物被膜的形成机制将有助于制定未来的控制策略。 霍乱流行，并有助于确定在感染期间对抗病原体的新药物靶标。

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.