Genetic Control of Growth on Surfaces

副溶血弧菌适应于表面的共生生活。当生长在表面或粘性层上时，这种有机体会诱导一个巨大的基因系统，并分化为沼泽细胞。向沼泽细胞的分化允许在表面和粘性环境中移动。细胞相互识别，运动本质上是协调的或社会的，导致有组织的社区中复杂的多细胞行为和增长。沼泽细胞有很好的移动和定植表面的能力：它是细长的和有超鞭毛的。这项工作的重点是揭开参与表面适应生活方式的调控网络，以便深入了解细菌如何在表面形成和维持群落。这个假设是，在这些群落的发展过程中，不同的基因组被开启和关闭。已知有一大组编码侧鞭毛运动系统的基因。第一个目标将是识别由表面生长诱导的基因；已经设计了一种新的转座子和递送系统来实现对表面诱导基因的搜索。这一目标的完成将导致识别尚未系统解剖的侧鞭毛层所涉及的基因，以及与表面适应的生活方式相关但不需要运动的新基因。将开发一张关于表面附着细菌的基因活动(基因鉴定和表达时序)的全面图片；这可能会揭示存在相互连接的调节电路来协调基因表达。第二个目标集中在最近发现的一种潜在电路上。两个连锁操纵子的突变会导致沼泽细胞基因的不可诱导表达。一个操纵子(Fim)编码一对菌毛状的伴侣-引座子和可能与外膜相关的蛋白质。第二个操纵子(Fli)编码一个ABC周质型结合蛋白(类似于大肠杆菌FliY)和一个类似于GGDEF-型传感器家族中的一些蛋白质的产物。我们将结合诱变、过表达和蛋白质定位实验来剖析这些基因组和游动细胞分化之间的相互作用。这些研究应该定义与表面生长相关的基因活性，并追踪使细菌能够在表面建立生长和发展群落的调控电路网络。了解表面诱导的基因表达模式应该揭示关于细菌如何适应环境、协调行为和发展有结构的群落或生物膜的重要原则。此外，该项目将对弧菌的遗传分析产生重大的教育影响，特别是转座子突变和全基因组搜索表面控制基因，将被用作本科生和研究生水平的教学工具。

V. parahaemolyticus is adapted to communal life on surfaces. When grown on a surface or in a viscous layer, the organism induces a large gene system and differentiates into a swarmer cell. Differentiation to the swarmer cell allows movement over surfaces and through viscous environments. Cells recognize each other, and movement is coordinated or social in nature, resulting in complex multicellular behavior and growth in organized communities. The swarmer cell is well-equipped to move over and colonize surfaces: it is elongated and hyperflagellated. The focus of this work is to unravel the regulatory networks that participate in the surface-adapted lifestyle in order to gain insight into how bacteria form and maintain communities on surfaces. The hypothesis is that different gene sets are turned on and off during the development of these communities. One large set, encoding the lateral flagellar motility system, is known. The first aim will be to identify genes induced by growth on surfaces; a novel transposon and delivery system has been designed to implement search for surface-induced genes. Completion of this aim will result in the identification of genes involved in the lateral flagellar hierarchy, which has not been systematically dissected, and new genes not required for motility but pertinent to the surface-adapted lifestyle. A comprehensive picture of gene activity (gene identity and timing of expression) of surface-attached bacteria will be developed; this will likely reveal the existence of interconnecting regulatory circuits to coordinate gene expression. The second aim focuses on one potential circuit that has recently been discovered. Mutations in two linked operons cause uninducible expression of swarmer cell genes. One operon (fim) encodes a fimbrial-like chaperone-usher pair and proteins that may be outer-membrane associated. The second operon (fli) encodes an ABC periplasmic-type binding protein (similar to Escherichia coli FliY) and a product that resembles a number of proteins in the GGDEF-type sensor family. The interactions between these gene sets and swarmer cell differentiation will be dissected by using a combination of mutagenesis, overexpression, and protein localization experiments. These studies should define gene activity pertinent to growth on surfaces and trace the network of regulatory circuitry that enables bacteria to establish growth and develop communities on surfaces. Understanding the pattern of surface-induced gene expression should reveal important principles about how bacteria adapt to their environment, coordinate behavior, and develop structured communities or biofilms. Moreover, this project will have significant educational impact for genetic analysis in Vibrio, specifically transposon mutagenesis and the genome-wide search for surface-controlled genes, will be used as an undergraduate and graduate level teaching tool.