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LuxU connects quorum sensing to biofilm formation in Vibrio fischeri.

ABSTRACT: Biofilm formation by Vibrio fischeri is a complex process involving multiple regulators, including the sensor kinase (SK) RscS and the response regulator (RR) SypG, which control the symbiosis polysaccharide (syp) locus. To identify other regulators of biofilm formation in V.?fischeri, we screened a transposon library for mutants defective in wrinkled colony formation. We identified LuxQ as a positive regulator of syp-dependent biofilm formation. LuxQ is a member of the Lux phosphorelay and is predicted to control bioluminescence in concert with the SK AinR, the phosphotransferase LuxU and the RR LuxO. Of these, LuxU was the only other regulator that exerted a substantial impact on biofilm formation. We propose a model in which the Lux pathway branches at LuxU to control both bioluminescence and biofilm formation. Furthermore, our evidence suggests that LuxU functions to regulate syp transcription, likely by controlling SypG activity. Finally, we found that, in contrast to its predicted function, the SK AinR has little impact on bioluminescence under our conditions. Thus, this study reveals a novel connection between the Lux and Syp pathways in V.?fischeri, and furthers our understanding of how the Lux pathway regulates bioluminescence in this organism.

SUBMITTER: Ray VA

PROVIDER: S-EPMC3566283 | biostudies-literature | 2012 Nov

REPOSITORIES: biostudies-literature

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LuxU connects quorum sensing to biofilm formation in Vibrio fischeri.

Ray Valerie A VA Visick Karen L KL

Molecular microbiology 20121005 4

Biofilm formation by Vibrio fischeri is a complex process involving multiple regulators, including the sensor kinase (SK) RscS and the response regulator (RR) SypG, which control the symbiosis polysaccharide (syp) locus. To identify other regulators of biofilm formation in V. fischeri, we screened a transposon library for mutants defective in wrinkled colony formation. We identified LuxQ as a positive regulator of syp-dependent biofilm formation. LuxQ is a member of the Lux phosphorelay and is p ...[more]

PMID: 23035866

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Project description:BACKGROUND: In a previous study, we demonstrated that Vibrio scophthalmi, the most abundant Vibrio species among the marine aerobic or facultatively anaerobic bacteria inhabiting the intestinal tract of healthy cultured turbot (Scophthalmus maximus), contains at least two quorum-sensing circuits involving two types of signal molecules (a 3-hydroxy-dodecanoyl-homoserine lactone and the universal autoinducer 2 encoded by luxS). The purpose of this study was to investigate the functions regulated by these quorum sensing circuits in this vibrio by constructing mutants for the genes involved in these circuits. RESULTS: The presence of a homologue to the Vibrio harveyi luxR gene encoding a main transcriptional regulator, whose expression is modulated by quorum-sensing signal molecules in other vibrios, was detected and sequenced. The V. scophthalmi LuxR protein displayed a maximum amino acid identity of 82% with SmcR, the LuxR homologue found in Vibrio vulnificus. luxR and luxS null mutants were constructed and their phenotype analysed. Both mutants displayed reduced biofilm formation in vitro as well as differences in membrane protein expression by mass-spectrometry analysis. Additionally, a recombinant strain of V. scophthalmi carrying the lactonase AiiA from Bacillus cereus, which causes hydrolysis of acyl homoserine lactones, was included in the study. CONCLUSIONS: V. scophthalmi shares two quorum sensing circuits, including the main transcriptional regulator luxR, with some pathogenic vibrios such as V. harveyi and V. anguillarum. However, contrary to these pathogenic vibrios no virulence factors (such as protease production) were found to be quorum sensing regulated in this bacterium. Noteworthy, biofilm formation was altered in luxS and luxR mutants. In these mutants a different expression profile of membrane proteins were observed with respect to the wild type strain suggesting that quorum sensing could play a role in the regulation of the adhesion mechanisms of this bacterium.

Project description:Biofilms, complex communities of microorganisms surrounded by a self-produced matrix, facilitate attachment and provide protection to bacteria. A natural model used to study biofilm formation is the symbiosis between Vibrio fischeri and its host, the Hawaiian bobtail squid, Euprymna scolopes Host-relevant biofilm formation is a tightly regulated process and is observed in vitro only with strains that have been genetically manipulated to overexpress or disrupt specific regulators, primarily two-component signaling (TCS) regulators. These regulators control biofilm formation by dictating the production of the symbiosis polysaccharide (Syp-PS), the major component of the biofilm matrix. Control occurs both at and below the level of transcription of the syp genes, which are responsible for Syp-PS production. Here, we probed the roles of the two known negative regulators of biofilm formation, BinK and SypE, by generating double mutants. We also mapped and evaluated a point mutation using natural transformation and linkage analysis. We examined traditional biofilm formation phenotypes and established a new assay for evaluating the start of biofilm formation in the form of microscopic aggregates in shaking liquid cultures, in the absence of the known biofilm-inducing signal calcium. We found that wrinkled colony formation is negatively controlled not only by BinK and SypE but also by SypF. SypF is both required for and inhibitory to biofilm formation. Together, these data reveal that these three regulators are sufficient to prevent wild-type V. fischeri from forming biofilms under these conditions.IMPORTANCE Bacterial biofilms promote attachment to a variety of surfaces and protect the constituent bacteria from environmental stresses, including antimicrobials. Understanding the mechanisms by which biofilms form will promote our ability to resolve them when they occur in the context of an infection. In this study, we found that Vibrio fischeri tightly controls biofilm formation using three negative regulators; the presence of a single one of these regulators was sufficient to prevent full biofilm development, while disruption of all three permitted robust biofilm formation. This work increases our understanding of the functions of specific regulators and demonstrates the substantial negative control that one benign microbe exerts over biofilm formation, potentially to ensure that it occurs only under the appropriate conditions.

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LuxU connects quorum sensing to biofilm formation in Vibrio fischeri.

Publications

LuxU connects quorum sensing to biofilm formation in Vibrio fischeri.

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