Analysis of surface behavior determinants in Variovorax paradoxus

描述（由申请人提供）：我们假设协调的表面行为，如群体运动和生物膜的形成，是在不同环境中竞争成功的关键。我们建议在一个模型系统Variovorax paradoxus中研究这些复杂表面行为的细节。根据我们实验室以前的工作，我们已经证明了在蜂群中产生的润湿剂的存在。我们将鉴定这种润湿剂和参与产生这种物质的基因。基于先前对悖论弧菌生物膜的研究，我们已经确定了IV型菌毛位点以及许多涉及影响群体和生物膜表型的脂多糖和外多糖的基因。我们将进一步研究pilY1基因的作用，该基因涉及表面行为的几个方面，并且可能存在于几种同种异构体中。我们的初步数据显示，该基因的独立插入突变可能具有不同的表型，这与两种蛋白质亚型在这些行为中发挥不同作用的假设一致。我们将研究LPS/eps合成位点的基因，并利用遗传和生化工具确定它们在合成表面和分泌多糖中的作用。我们将研究控制这些表面行为之间决定的培养条件，并识别在群体运动中有缺陷的突变体，重点关注触发反应的信号转导途径。这些实验将使我们能够建立一个悖论弧菌表面行为的模型，其中包括对运动和无根发育至关重要的结构成分，以及控制这一决定的信号转导途径和生化回路。

DESCRIPTION (provided by applicant): We hypothesize that coordinated surface behavior, such as swarming motility and biofilm formation, is crucial for competitive success in diverse environments. We propose to examine the details of these complex surface behaviors in a model system, Variovorax paradoxus. Based on previous work in our lab, we have demonstrated the presence of a wetting agent that is produced during swarming. We will identify this wetting agent and the genes involved in producing the material. Based on previous work on V. paradoxus biofilms, we have identified a type IV pilus locus as well as numerous genes involved in lipopolysaccharide and exopolysaccharides that influence swarming and biofilm phenotypes. We will further examine the roles of the pilY1 gene, which has been implicated in several aspects of surface behavior, and which may be present in several isoforms. Our preliminary data shows that independent insertion mutants in this gene can have divergent phenotypes, consistent with the hypothesis that two protein isoforms are playing different roles in these behaviors. The genes of the LPS/eps synthesis loci will be investigated and their roles synthesis of surface and secreted polysaccharides will be determined using genetic and biochemical tools. We will examine the culture conditions that control the decision between these surface behaviors, and identify mutants that are defective in swarming motility, focusing on the signal transduction pathways that trigger the response. These experiments will allow us to build a model of V. paradoxus surface behaviors that includes the structural components critical to motility and sessile development, as well as the signal transduction pathways and biochemical circuits that control this decision. PUBLIC HEALTH RELEVANCE: Bacterial success in many environments is dependent on coordinated surface behaviors, including motility and sessile development as a biofilm. In this study, we examine the surface behaviors and associated structures -- such as wetting agents, pili, flagella, and exopolysaccharides-- of the soil bacterium Variovorax paradoxus, the genes involved in these behaviors and their regulation by environmental cues. These studies will result in a comprehensive model of these surface behaviors, which are crucial to bacterial survival in many environments, with particular relevance to opportunistic infections and virulence.