Articoli di riviste: "LuxR"

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Bazhenov, Sergey, Olga Melkina, Vadim Fomin, Ekaterina Scheglova, Pavel Krasnik, Svetlana Khrulnova, Gennadii Zavilgelsky e Ilya Manukhov. "LitR directly upregulates autoinducer synthesis and luminescence in Aliivibrio logei". PeerJ 9 (21 settembre 2021): e12030. http://dx.doi.org/10.7717/peerj.12030.

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LitR is a master-regulator of transcription in the ainS/R and luxS/PQ quorum sensing (QS) systems of bacteria from Vibrio and Aliivibrio genera. Here, we for the first time directly investigated the influence of LitR on gene expression in the luxI/R QS system of psychrophilic bacteria Aliivibrio logei. Investigated promoters were fused with Photorhabdus luminescens luxCDABE reporter genes cassette in a heterological system of Escherichia coli cells, litR A. logei was introduced into the cells under control of Plac promoter. LitR has been shown to upregulate genes of autoinducer synthase (luxI), luciferase and reductase (luxCDABE), and this effect doesn’t depend on presence of luxR gene. To a much lesser degree, LitR induces luxR1, but not the luxR2 — the main luxI/R regulator. Enhanced litR expression leads to an increase in a LuxI-autoinducer synthesis and a subsequent LuxR-mediated activation of the luxI/R QS system. Effect of LitR on luxI transcription depends on lux-box sequence in luxI promoter even in absence of luxR (lux-box is binding site of LuxR). The last finding indicates a direct interaction of LitR with the promoter in the lux-box region. Investigation of the effect of LitR A. logei on luxI/R QS systems of mesophilic Aliivibrio fischeri and psychrophilic Aliivibrio salmonicida showed direct luxR-independent upregulation of luxI and luxCDABE genes. To a lesser degree, it induces luxR A. fischeri and luxR1 A. salmonicida. Therefore, we assume that the main role of LitR in cross-interaction of these three QS systems is stimulating the expression of luxI.

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Bazhenov, Sergey, Uliana Novoyatlova, Ekaterina Scheglova, Vadim Fomin, Svetlana Khrulnova, Olga Melkina, Vladimir Chistyakov e Ilya Manukhov. "Influence of the luxR Regulatory Gene Dosage and Expression Level on the Sensitivity of the Whole-Cell Biosensor to Acyl-Homoserine Lactone". Biosensors 11, n. 6 (23 maggio 2021): 166. http://dx.doi.org/10.3390/bios11060166.

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Aliivibrio fischeri LuxR and Aliivibrio logei LuxR1 and LuxR2 regulatory proteins are quorum sensing transcriptional (QS) activators, inducing promoters of luxICDABEG genes in the presence of an autoinducer (3-oxo-hexanoyl-l-homoserine lactone). In the Aliivibrio cells, luxR genes are regulated by HNS, CRP, LitR, etc. Here we investigated the role of the luxR expression level in LuxI/R QS system functionality and improved the whole-cell biosensor for autoinducer detection. Escherichia coli-based bacterial lux-biosensors were used, in which Photorhabdus luminescensluxCDABE genes were controlled by LuxR-dependent promoters and luxR, luxR1, or luxR2 regulatory genes. We varied either the dosage of the regulatory gene in the cells using additional plasmids, or the level of the regulatory gene expression using the lactose operon promoter. It was shown that an increase in expression level, as well as dosage of the regulatory gene in biosensor cells, leads to an increase in sensitivity (the threshold concentration of AI is reduced by one order of magnitude) and to a two to threefold reduction in response time. The best parameters were obtained for a biosensor with an increased dosage of luxRA. fischeri (sensitivity to 3-oxo-hexanoyl-l-homoserine lactone reached 30–100 pM).

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Urbanowski, M. L., C. P. Lostroh e E. P. Greenberg. "Reversible Acyl-Homoserine Lactone Binding to Purified Vibrio fischeri LuxR Protein". Journal of Bacteriology 186, n. 3 (1 febbraio 2004): 631–37. http://dx.doi.org/10.1128/jb.186.3.631-637.2004.

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ABSTRACT The Vibrio fischeri LuxR protein is the founding member of a family of acyl-homoserine lactone-responsive quorum-sensing transcription factors. Previous genetic evidence indicates that in the presence of its quorum-sensing signal, N-(3-oxohexanoyl) homoserine lactone (3OC6-HSL), LuxR binds to lux box DNA within the promoter region of the luxI gene and activates transcription of the luxICDABEG luminescence operon. We have purified LuxR from recombinant Escherichia coli. Purified LuxR binds specifically and with high affinity to DNA containing a lux box. This binding requires addition of 3OC6-HSL to the assay reactions, presumably forming a LuxR-3OC6-HSL complex. When bound to the lux box at the luxI promoter in vitro, LuxR-3OC6-HSL enables E. coli RNA polymerase to initiate transcription from the luxI promoter. Unlike the well-characterized LuxR homolog TraR in complex with its signal (3-oxo-octanoyl-HSL), the LuxR-30C6-HSL complex can be reversibly inactivated by dilution, suggesting that 3OC6-HSL in the complex is not tightly bound and is in equilibrium with the bulk solvent. Thus, although LuxR and TraR both bind 3-oxoacyl-HSLs, the binding is qualitatively different. The differences have implications for the ways in which these proteins respond to decreases in signal concentrations or rapid drops in population density.

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Antunes, Luis Caetano M., Rosana B. R. Ferreira, C. Phoebe Lostroh e E. Peter Greenberg. "A Mutational Analysis Defines Vibrio fischeri LuxR Binding Sites". Journal of Bacteriology 190, n. 13 (14 dicembre 2007): 4392–97. http://dx.doi.org/10.1128/jb.01443-07.

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ABSTRACT Vibrio fischeri quorum sensing involves the LuxI and LuxR proteins. The LuxI protein generates the quorum-sensing signal N-3-oxohexanoyl-l-homoserine lactone (3OC6-HSL), and LuxR is a signal-responsive transcriptional regulator which activates the luminescence (lux) genes and 17 other V. fischeri genes. For activation of the lux genes, LuxR binds to a 20-base-pair inverted repeat, the lux box, which is centered 42.5 base pairs upstream of the transcriptional start of the lux operon. Similar lux box-like elements have been identified in only a few of the LuxR-activated V. fischeri promoters. To better understand the DNA sequence elements required for LuxR binding and to identify binding sites in LuxR-regulated promoters other than the lux operon promoter, we have systematically mutagenized the lux box and evaluated the activity of many mutants. By doing so, we have identified nucleotides that are critical for promoter activity. Interestingly, certain lux box mutations allow a 3OC6-HSL-independent LuxR activation of the lux operon promoter. We have used the results of the mutational analysis to create a consensus lux box, and we have used this consensus sequence to identify LuxR binding sites in 3OC6-HSL-activated genes for which lux boxes could not be identified previously.

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Swearingen, Matthew C., Anice Sabag-Daigle e Brian M. M. Ahmer. "Are There Acyl-Homoserine Lactones within Mammalian Intestines?" Journal of Bacteriology 195, n. 2 (9 novembre 2012): 173–79. http://dx.doi.org/10.1128/jb.01341-12.

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ABSTRACTManyProteobacteriaare capable of quorum sensing usingN-acyl-homoserine lactone (acyl-HSL) signaling molecules that are synthesized by LuxI or LuxM homologs and detected by transcription factors of the LuxR family. Most quorum-sensing species have at least one LuxR and one LuxI homolog. However, members of theEscherichia,Salmonella,Klebsiella, andEnterobactergenera possess only a single LuxR homolog, SdiA, and no acyl-HSL synthase. The most obvious hypothesis is that these organisms are eavesdropping on acyl-HSL production within the complex microbial communities of the mammalian intestinal tract. However, there is currently no evidence of acyl-HSLs being produced within normal intestinal communities. A few intestinal pathogens, includingYersinia enterocolitica, do produce acyl-HSLs, andSalmonellacan detect them during infection. Therefore, a more refined hypothesis is that SdiA orthologs are used for eavesdropping on other quorum-sensing pathogens in the host. However, the lack of acyl-HSL signaling among the normal intestinal residents is a surprising finding given the complexity of intestinal communities. In this review, we examine the evidence for and against the possibility of acyl-HSL signaling molecules in the mammalian intestine and discuss the possibility that related signaling molecules might be present and awaiting discovery.

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Antunes, Luis Caetano M., Amy L. Schaefer, Rosana B. R. Ferreira, Nan Qin, Ann M. Stevens, Edward G. Ruby e E. Peter Greenberg. "Transcriptome Analysis of the Vibrio fischeri LuxR-LuxI Regulon". Journal of Bacteriology 189, n. 22 (7 settembre 2007): 8387–91. http://dx.doi.org/10.1128/jb.00736-07.

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ABSTRACT The Vibrio fischeri quorum-sensing signal N-3-oxohexanoyl-l-homoserine lactone (3OC6-HSL) activates expression of the seven-gene luminescence operon. We used microarrays to unveil 18 additional 3OC6-HSL-controlled genes, 3 of which had been identified by other means previously. We show most of these genes are regulated by the 3OC6-HSL-responsive transcriptional regulator LuxR directly. This demonstrates that V. fischeri quorum sensing regulates a substantial number of genes other than those involved in light production.

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von Bodman, Susanne B., Jessica K. Ball, Marie A. Faini, Carmen M. Herrera, Timothy D. Minogue, Mark L. Urbanowski e Ann M. Stevens. "The Quorum Sensing Negative Regulators EsaR and ExpREcc, Homologues within the LuxR Family, Retain the Ability To Function as Activators of Transcription". Journal of Bacteriology 185, n. 23 (1 dicembre 2003): 7001–7. http://dx.doi.org/10.1128/jb.185.23.7001-7007.2003.

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ABSTRACT Most LuxR homologues function as activators of transcription during the process of quorum sensing, but a few, including EsaR and ExpR Ecc , negatively impact gene expression. The LuxR-activated luxI promoter and LuxR binding site, the lux box, were used in artificial contexts to assess the potential for transcriptional activation and DNA binding by EsaR and ExpR Ecc . Although the acyl-homoserine lactone responsiveness of both proteins is the opposite of that shown by most LuxR family members, EsaR and ExpR Ecc have preserved the ability to interact with RNA polymerase and activate transcription despite their low affinity for the lux box DNA.

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Subramoni, Sujatha, e Vittorio Venturi. "LuxR-family ‘solos’: bachelor sensors/regulators of signalling molecules". Microbiology 155, n. 5 (1 maggio 2009): 1377–85. http://dx.doi.org/10.1099/mic.0.026849-0.

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N-Acylhomoserine lactone (AHL) quorum-sensing (QS) signalling is the best-understood chemical language in proteobacteria. In the last 15 years a large amount of research in several bacterial species has revealed in detail the genetic, molecular and biochemical mechanisms underlying AHL signalling. These studies have revealed the role played by protein pairs of the AHL synthase belonging to the LuxI family and cognate LuxR-family AHL sensor–regulator. Proteobacteria however commonly possess a QS LuxR-family protein for which there is no obvious cognate LuxI synthase; these proteins are found in bacteria which possess a complete AHL QS system(s) as well as in bacteria that do not. Scientists are beginning to address the roles played by these proteins and it is emerging that they could allow bacteria to respond to endogenous and exogenous signals produced by their neighbours. AHL QS research thus far has mainly focused on a cell-density response involving laboratory monoculture studies. Recent findings on the role played by the unpaired LuxR-family proteins highlight the need to address bacterial behaviour and response to signals in mixed communities. Here we review recent progress with respect to these LuxR proteins, which we propose to call LuxR ‘solos’ since they act on their own without the need for a cognate signal generator. Initial investigations have revealed that LuxR solos have diverse roles in bacterial interspecies and interkingdom communication.

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McDougald, Diane, Scott A. Rice e Staffan Kjelleberg. "SmcR-Dependent Regulation of Adaptive Phenotypes inVibrio vulnificus". Journal of Bacteriology 183, n. 2 (15 gennaio 2001): 758–62. http://dx.doi.org/10.1128/jb.183.2.758-762.2001.

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ABSTRACT Vibrio vulnificus contains homologues of the V. harveyi luxR and luxS genes. A null mutation insmcR (luxR) resulted in a defect in starvation survival, inhibition of starvation-induced maintenance of culturability that occurs when V. vulnificusis starved prior to low-temperature incubation, and increased expression of stationary-phase phenotypes.

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Gray, Kendall M., e James R. Garey. "The evolution of bacterial LuxI and LuxR quorum sensing regulators". Microbiology 147, n. 8 (1 agosto 2001): 2379–87. http://dx.doi.org/10.1099/00221287-147-8-2379.

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Brameyer, Sophie, Darko Kresovic, Helge B. Bode e Ralf Heermann. "Dialkylresorcinols as bacterial signaling molecules". Proceedings of the National Academy of Sciences 112, n. 2 (30 dicembre 2014): 572–77. http://dx.doi.org/10.1073/pnas.1417685112.

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It is well recognized that bacteria communicate via small diffusible molecules, a process termed quorum sensing. The best understood quorum sensing systems are those that use acylated homoserine lactones (AHLs) for communication. The prototype of those systems consists of a LuxI-like AHL synthase and a cognate LuxR receptor that detects the signal. However, many proteobacteria possess LuxR receptors, yet lack any LuxI-type synthase, and thus these receptors are referred to as LuxR orphans or solos. In addition to the well-known AHLs, little is known about the signaling molecules that are sensed by LuxR solos. Here, we describe a novel cell–cell communication system in the insect and human pathogenPhotorhabdus asymbiotica. We identified the LuxR homolog PauR to sense dialkylresorcinols (DARs) and cyclohexanediones (CHDs) instead of AHLs as signals. The DarABC synthesis pathway produces the molecules, and the entire system emerged as important for virulence. Moreover, we have analyzed more than 90 differentPhotorhabdusstrains by HPLC/MS and showed that these DARs and CHDs are specific to the human pathogenP. asymbiotica. On the basis of genomic evidence, 116 other bacterial species are putative DAR producers, among them many human pathogens. Therefore, we discuss the possibility of DARs as novel and widespread bacterial signaling molecules and show that bacterial cell–cell communication goes far beyond AHL signaling in nature.

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Zheng, Huiming, Yiling Mao, Qingcheng Zhu, Jun Ling, Na Zhang, Nawar Naseer, Zengtao Zhong e Jun Zhu. "The Quorum Sensing Regulator CinR Hierarchically Regulates Two Other Quorum Sensing Pathways in Ligand-Dependent and -Independent Fashions in Rhizobium etli". Journal of Bacteriology 197, n. 9 (17 febbraio 2015): 1573–81. http://dx.doi.org/10.1128/jb.00003-15.

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ABSTRACTMany rhizobial species use complexN-acyl-homoserine lactone (AHL)-based quorum sensing (QS) systems to monitor their population density and regulate their symbiotic interactions with their plant hosts. There are at least three LuxRI-type regulatory systems inRhizobium etliCFN42: CinRI, RaiRI, and TraRI. In this study, we show that CinI, RaiI, and TraI are responsible for synthesizing all AHLs under the tested conditions. The activation of these AHL synthase genes requires their corresponding LuxR-type counterparts. We further demonstrate that CinRI is at the top of the regulatory cascade that activates RaiRI and TraRI QS systems. Moreover, we discovered that CinR possesses a specific affinity to bindcinIpromoter in the absence of its cognate AHL ligand, thereby activatingcinItranscription. Addition of AHLs leads to improved binding to thecinIpromoter and enhancedcinIexpression. Furthermore, we found that compared to the wild type, thecinRmutation displayed reduced nodule formation, andcinR,raiR, andtraImutants show significantly lower levels of nitrogen fixation activity than the wild type. These results suggest that the complex QS regulatory systems inR. etliplay an important role in its symbiosis with legume hosts.IMPORTANCEMany bacteria use quorum sensing (QS) to monitor their cell densities and coordinately regulate a number of physiological functions. Rhizobia often have diverse and complex LuxR/LuxI-type quorum sensing systems that may be involved in symbiosis and N2fixation. In this study, we identified three LuxR/LuxI-type QS systems inRhizobium etliCFN42: CinRI, RaiRI, and TraRI. We established a complex network of regulation between these QS components and found that these QS systems played important roles in symbiosis processes.

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Truong, Thao T., Mohammad Seyedsayamdost, E. Peter Greenberg e Josephine R. Chandler. "A Burkholderia thailandensis Acyl-Homoserine Lactone-Independent Orphan LuxR Homolog That Activates Production of the Cytotoxin Malleilactone". Journal of Bacteriology 197, n. 21 (17 agosto 2015): 3456–62. http://dx.doi.org/10.1128/jb.00425-15.

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ABSTRACTBurkholderia thailandensishas three acyl-homoserine lactone (AHL) LuxR-LuxI quorum-sensing circuits and two orphan LuxR homologs. Orphans are LuxR-type transcription factors that do not have cognate LuxI-type AHL synthases. One of the orphans, MalR, is genetically linked to themalgene cluster, which encodes enzymes required for production of the cytotoxic polyketide malleilactone. Under normal laboratory conditions themalgene cluster is silent; however, antibiotics like trimethoprim inducemaltranscription. We show that trimethoprim-dependent induction of themalgenes requires MalR. MalR has all of the conserved amino acid residues characteristic of AHL-responsive LuxR homologs, but inB. thailandensis, MalR activation of malleilactone synthesis genes is not responsive to AHLs. MalR can activate transcription from themalpromoter inE. coliwithout addition of AHLs or trimethoprim. Expression ofmalRinB. thailandensisis induced by trimethoprim. Our data indicate that MalR binds to aluxbox-like element in themalpromoter and activates transcription of themalgenes in an AHL-independent manner. Antibiotics like trimethoprim appear to activatemalgene expression indirectly by somehow activatingmalRexpression. MalR activation of themalgenes represents an example of a LuxR homolog that is not a receptor for an AHL quorum-sensing signal. Our evidence is consistent with the idea thatmalgene activation depends solely on sufficient transcription of themalRgene.IMPORTANCELuxR proteins are transcription factors that are typically activated by acyl-homoserine lactone (AHL) signals. We demonstrate that a conserved LuxR family protein, MalR, activates genes independently of AHLs. MalR is required for transcription of genes coding for synthesis of the cytotoxic polyketide malleilactone. These genes are not expressed when cells are grown under normal laboratory conditions. In laboratory culture, MalR induction of malleilactone requires certain antibiotics, such as trimethoprim, which increasemalRexpression by an unknown mechanism. At sufficient levels ofmalRexpression, MalR functions independently of any external signal. Our findings show that MalR is an activator of the silent malleilactone biosynthesis genes and that MalR functions independently of AHLs.

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Nasuno, Eri, Nobutada Kimura, Masaki J. Fujita, Cindy H. Nakatsu, Yoichi Kamagata e Satoshi Hanada. "Phylogenetically Novel LuxI/LuxR-Type Quorum Sensing Systems Isolated Using a Metagenomic Approach". Applied and Environmental Microbiology 78, n. 22 (14 settembre 2012): 8067–74. http://dx.doi.org/10.1128/aem.01442-12.

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ABSTRACTA great deal of research has been done to understand bacterial cell-to-cell signaling systems, but there is still a large gap in our current knowledge because the majority of microorganisms in natural environments do not have cultivated representatives. Metagenomics is one approach to identify novel quorum sensing (QS) systems from uncultured bacteria in environmental samples. In this study, fosmid metagenomic libraries were constructed from a forest soil and an activated sludge from a coke plant, and the target genes were detected using a green fluorescent protein (GFP)-basedEscherichia colibiosensor strain whose fluorescence was screened by spectrophotometry. DNA sequence analysis revealed two pairs of new LuxI familyN-acyl-l-homoserine lactone (AHL) synthases and LuxR family transcriptional regulators (clones N16 and N52, designated AubI/AubR and AusI/AusR, respectively). AubI and AusI each produced an identical AHL,N-dodecanoyl-l-homoserine lactone (C12-HSL), as determined by nuclear magnetic resonance (NMR) and mass spectrometry. Phylogenetic analysis based on amino acid sequences suggested that AusI/AusR was from an uncultured member of theBetaproteobacteriaand AubI/AubR was very deeply branched from previously described LuxI/LuxR homologues in isolates of theProteobacteria. The phylogenetic position of AubI/AubR indicates that they represent a QS system not acquired recently from theProteobacteriaby horizontal gene transfer but share a more ancient ancestry. We demonstrated that metagenomic screening is useful to provide further insight into the phylogenetic diversity of bacterial QS systems by describing two new LuxI/LuxR-type QS systems from uncultured bacteria.

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Plener, Laure, Nicola Lorenz, Matthias Reiger, Tiago Ramalho, Ulrich Gerland e Kirsten Jung. "The Phosphorylation Flow of the Vibrio harveyi Quorum-Sensing Cascade Determines Levels of Phenotypic Heterogeneity in the Population". Journal of Bacteriology 197, n. 10 (9 marzo 2015): 1747–56. http://dx.doi.org/10.1128/jb.02544-14.

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ABSTRACTQuorum sensing (QS) is a communication process that enables a bacterial population to coordinate and synchronize specific behaviors. The bioluminescent marine bacteriumVibrio harveyiintegrates three autoinducer (AI) signals into one quorum-sensing cascade comprising a phosphorelay involving three hybrid sensor kinases: LuxU; LuxO, an Hfq/small RNA (sRNA) switch; and the transcriptional regulator LuxR. Using a new set ofV. harveyimutants lacking genes for the AI synthases and/or sensors, we assayed the activity of the quorum-sensing cascade at the population and single-cell levels, with a specific focus on signal integration and noise levels. We found that the ratios of kinase activities to phosphatase activities of the three sensors and, hence, the extent of phosphorylation of LuxU/LuxO are important not only for the signaling output but also for the degree of noise in the system. The pools of phosphorylated LuxU/LuxO per cell directly determine the amounts of sRNAs produced and, consequently, the copy number of LuxR, generating heterogeneous quorum-sensing activation at the single-cell level. We conclude that the ability to drive the heterogeneous expression of QS-regulated genes inV. harveyiis an inherent feature of the architecture of the QS cascade.IMPORTANCEV. harveyipossesses one of the most complex quorum-sensing (QS) cascades known, using three different autoinducers (AIs) to control the induction of, e.g., bioluminescence, virulence factors, and biofilm and exoprotease production. We constructed variousV. harveyimutants to study the impact of each component and subsystem of the QS signaling cascade on QS activation at the population and single-cell levels. We found that the output was homogeneous only in the presence of all AIs. In the absence of any one AI, QS activation varied from cell to cell, resulting in phenotypic heterogeneity. This study elucidates a molecular design principle which enables a tightly integrated signaling cascade to control the expression of diverse phenotypes within a genetically homogeneous population.

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Rutherford, Steven T., Julie S. Valastyan, Thibaud Taillefumier, Ned S. Wingreen e Bonnie L. Bassler. "Comprehensive analysis reveals how single nucleotides contribute to noncoding RNA function in bacterial quorum sensing". Proceedings of the National Academy of Sciences 112, n. 44 (19 ottobre 2015): E6038—E6047. http://dx.doi.org/10.1073/pnas.1518958112.

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Five homologous noncoding small RNAs (sRNAs), called the Qrr1-5 sRNAs, function in the Vibrio harveyi quorum-sensing cascade to drive its operation. Qrr1-5 use four different regulatory mechanisms to control the expression of ∼20 mRNA targets. Little is known about the roles individual nucleotides play in mRNA target selection, in determining regulatory mechanism, or in defining Qrr potency and dynamics of target regulation. To identify the nucleotides vital for Qrr function, we developed a method we call RSort-Seq that combines saturating mutagenesis, fluorescence-activated cell sorting, high-throughput sequencing, and mutual information theory to explore the role that every nucleotide in Qrr4 plays in regulation of two mRNA targets, luxR and luxO. Companion biochemical assays allowed us to assign specific regulatory functions/underlying molecular mechanisms to each important base. This strategy yielded a regional map of nucleotides in Qrr4 vital for stability, Hfq interaction, stem-loop formation, and base pairing to both luxR and luxO, to luxR only, and to luxO only. In terms of nucleotides critical for sRNA function, the RSort-Seq analysis provided strikingly different results from those predicted by commonly used regulatory RNA-folding algorithms. This approach is applicable to any RNA–RNA interaction, including sRNAs in other bacteria and regulatory RNAs in higher organisms.

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White, Catharine E., e Stephen C. Winans. "Cell–cell communication in the plant pathogen Agrobacterium tumefaciens". Philosophical Transactions of the Royal Society B: Biological Sciences 362, n. 1483 (13 marzo 2007): 1135–48. http://dx.doi.org/10.1098/rstb.2007.2040.

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The plant pathogen Agrobacterium tumefaciens induces the formation of crown gall tumours at wound sites on host plants by directly transforming plant cells. This disease strategy benefits the bacteria as the infected plant tissue produces novel nutrients, called opines, that the colonizing bacteria can use as nutrients. Almost all of the genes that are required for virulence, and all of the opine uptake and utilization genes, are carried on large tumour-inducing (Ti) plasmids. The observation more than 25 years ago that specific opines are required for Ti plasmid conjugal transfer led to the discovery of a cell–cell signalling system on these plasmids that is similar to the LuxR–LuxI system first described in Vibrio fischeri . All Ti plasmids that have been described to date carry a functional LuxI-type N -acylhomoserine lactone synthase (TraI), and a LuxR-type signal receptor and transcriptional regulator called TraR. The traR genes are expressed only in the presence of specific opines called conjugal opines. The TraR–TraI system provides an important model for LuxR–LuxI-type systems, especially those found in the agriculturally important Rhizobiaceae family. In this review, we discuss current advances in the biochemistry and structural biology of the TraR–TraI system.

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Ulrich, Ricky L., David DeShazer, Harry B. Hines e Jeffrey A. Jeddeloh. "Quorum Sensing: a Transcriptional Regulatory System Involved in the Pathogenicity of Burkholderia mallei". Infection and Immunity 72, n. 11 (novembre 2004): 6589–96. http://dx.doi.org/10.1128/iai.72.11.6589-6596.2004.

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ABSTRACT Numerous gram-negative bacterial pathogens regulate virulence factor expression by using a cell density mechanism termed quorum sensing (QS). An in silico analysis of the Burkholderia mallei ATCC 23344 genome revealed that it encodes at least two luxI and four luxR homologues. Using mass spectrometry, we showed that wild-type B. mallei produces the signaling molecules N-octanoyl-homoserine lactone and N-decanoyl-homoserine lactone. To determine if QS is involved in the virulence of B. mallei, we generated mutations in each putative luxIR homologue and tested the pathogenicities of the derivative strains in aerosol BALB/c mouse and intraperitoneal hamster models. Disruption of the B. mallei QS alleles, especially in RJ16 (bmaII) and RJ17 (bmaI3), which are luxI mutants, significantly reduced virulence, as indicated by the survival of mice who were aerosolized with 104 CFU (10 50% lethal doses [LD50s]). For the B. mallei transcriptional regulator mutants (luxR homologues), mutation of the bmaR5 allele resulted in the most pronounced decrease in virulence, with 100% of the challenged animals surviving a dose of 10 LD50s. Using a Syrian hamster intraperitoneal model of infection, we determined the LD50s for wild-type B. mallei and each QS mutant. An increase in the relative LD50 was found for RJ16 (bmaI1) (>967 CFU), RJ17 (bmaI3) (115 CFU), and RJ20 (bmaR5) (151 CFU) compared to wild-type B. mallei (<13 CFU). These findings demonstrate that B. mallei carries multiple luxIR homologues that either directly or indirectly regulate the biosynthesis of an essential virulence factor(s) that contributes to the pathogenicity of B. mallei in vivo.

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Hawkins, Andrew C., Frances H. Arnold, Rainer Stuermer, Bernhard Hauer e Jared R. Leadbetter. "Directed Evolution of Vibrio fischeri LuxR for Improved Response to Butanoyl-Homoserine Lactone". Applied and Environmental Microbiology 73, n. 18 (3 agosto 2007): 5775–81. http://dx.doi.org/10.1128/aem.00060-07.

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ABSTRACT LuxR is the 3-oxohexanoyl-homoserine lactone (3OC6HSL)-dependent transcriptional activator of the prototypical acyl-homoserine lactone (AHL) quorum-sensing system of Vibrio fischeri. Wild-type LuxR exhibits no response to butanoyl-HSL (C4HSL) in quantitative bioassays at concentrations of up to 1 μM; a previously described LuxR variant (LuxR-G2E) exhibits a broadened response to diverse AHLs, including pentanoyl-HSL (C5HSL), but not to C4HSL. Here, two rounds of directed evolution of LuxR-G2E generated variants of LuxR that responded to C4HSL at concentrations as low as 10 nM. One variant, LuxR-G4E, had only one change, I45F, relative to the parent LuxR-G2E, which itself differs from the wild type at three residues. Dissection of the four mutations within LuxR-G4E demonstrated that at least three of these changes were simultaneously required to achieve any measurable C4HSL response. The four changes improved both sensitivity and specificity towards C4HSL relative to any of the other 14 possible combinations of those residues. These data confirm that LuxR is evolutionarily pliable and suggest that LuxR is not intrinsically asymmetric in its response to quorum-sensing signals with different acyl-side-chain lengths.

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Perry, Lynda L., Nathan G. Bright, Richard J. Carroll, Jr., M. Cathy Scott, Michael S. Allen e Bruce M. Applegate. "Molecular characterization of autoinduction of bioluminescence in the Microtox® indicator strain Vibrio fischeri ATCC 49387". Canadian Journal of Microbiology 51, n. 7 (1 luglio 2005): 549–57. http://dx.doi.org/10.1139/w05-019.

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Repeated attempts to clone the luxI from Vibrio fischeri ATCC 49387 failed to produce a clone carrying a functional LuxI. Sequence data from the clones revealed the presence of a polymorphism when compared with previously published luxI sequences, prompting further characterization of bioluminescence regulation in V. fischeri ATCC 49387. Further investigation of V. fischeri ATCC 49387 revealed that its LuxI protein lacks detectable LuxI activity due to the presence of a glutamine residue at position 125 in the deduced amino acid sequence. Specific bioluminescence in V. fischeri ATCC 49387 increases with increasing cell density, indicative of a typical autoinduction response. However, conditioned medium from this strain does not induce bioluminescence in an ATCC 49387 luxR-plux-based acyl homoserine lactone reporter strain, but does induce bioluminescence in ATCC 49387. It has been previously shown that a V. fischeri MJ-1 luxI mutant exhibits autoinduction of bioluminescence through N-octanoyl-L-homoserine lactone, the product of the AinS autoinducer synthase. However, a bioreporter based on luxR-plux from V. fischeri ATCC 49387 responded poorly to conditioned medium from V. fischeri ATCC 49387 and also responded poorly to authentic N-octanoyl-DL-homoserine lactone. A similar MJ-1-based bioreporter showed significant induction under the same conditions. A putative ainS gene cloned from ATCC 49387, unlike luxI from ATCC 49387, expresses V. fischeri autoinducer synthase activity in Escherichia coli. This study suggests that a regulatory mechanism independent of LuxR and LuxI but possibly involving AinS is responsible for the control of autoinduction of bioluminescence in V. fischeri ATCC 49387.Key words: quorum sensing, bioluminescence, Vibrio fischeri.

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Schu, Daniel J., Aurelien L. Carlier, Katherine P. Jamison, Susanne von Bodman e Ann M. Stevens. "Structure/Function Analysis of the Pantoea stewartii Quorum-Sensing Regulator EsaR as an Activator of Transcription". Journal of Bacteriology 191, n. 24 (9 ottobre 2009): 7402–9. http://dx.doi.org/10.1128/jb.00994-09.

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ABSTRACT In Pantoea stewartii subsp. stewartii, two regulatory proteins are key to the process of cell-cell communication known as quorum sensing: the LuxI and LuxR homologues EsaI and EsaR. Most LuxR homologues function as activators of transcription in the presence of their cognate acylated homoserine lactone (AHL) signal. However, EsaR was initially found to function as a repressor in the absence of AHL. Previous studies demonstrated that, in the absence of AHL, EsaR retains the ability to function as a weak activator of the lux operon in recombinant Escherichia coli. Here it is shown that both the N-terminal and the C-terminal domains of EsaR are necessary for positive regulation. A site-directed mutagenesis study, guided by homology modeling to LuxR and TraR, has revealed three critical residues in EsaR that are involved in activation of RNA polymerase. In addition, a native EsaR-activated promoter has been identified, which controls expression of a putative regulatory sRNA in P. stewartii.

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Grellier, N., M. Suzuki, L. Brot, A. Rodrigues, L. Humbert, K. Escoubeyrou, D. Rainteau, J. P. Grill, R. Lami e P. Seksik. "DOP50 Impact of Inflammatory Bowel Disease associated dysbiosis on bacterial quorum sensing mediated by Acyl-homoserine Lactone in human gut microbiota". Journal of Crohn's and Colitis 17, Supplement_1 (30 gennaio 2023): i119—i121. http://dx.doi.org/10.1093/ecco-jcc/jjac190.0090.

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Abstract Background Intestinal dysbiosis is a key feature in the pathogenesis of inflammatory bowel diseases (IBD). Bacterial quorum sensing mediated by acyl-homoserine lactones (AHL) might play a role in the dialogue between the gut microbiota and the host. The main objective of our study was to investigate the presence and expression of AHL synthase and receptor genes in the human gut ecosystem during IBD. Methods To confirm the presence of AHL in the gut, mass spectrometric detection was performed on stool samples from IBD patients and non-IBD subjects. Then, by an in silico approach, we exploited the open access database: Inflammatory Bowel Disease Multi'omics Database, an American cohort with bacterial metagenomes and metatranscriptomes data of stool samples from non-IBD and IBD subjects. To characterise gut dysbiosis, the most discriminating bacterial species between non-IBD and IBD patients were identified by multivariate analysis and allowed us to define two groups (dysbiosis/non-dysbiosis). The search for AHL synthase (luxI) and receptor (luxR) known homolog genes, was performed using Basic Local Alignment Search Tool (BLAST) from previously assembled gene files (presence/absence) as well as raw data sequencing files (relative abundance and expression). Results Mass spectrometry confirmed a higher concentration of AHL molecules in healthy subjects than in relapsing IBD. Regarding in silico analysis, 103 subjects were selected including 50 with Crohn's disease (CD), 27 with ulcerative colitis (UC), and 26 non-IBD subjects. No luxI-like synthase genes were retrieved by BLAST searches. However, several homologs of receptor genes were identified: sdiA gene from Escherichia coli (7/103 patients) and luxR-like homologs from Bacteroides fragilis and Bacteroides dorei present in all patients. According to disease, only one luxR-like gene from Bacteroides dorei was under-expressed in IBD patients (p = 0.02) compared to non-IBD, especially in CD (p = 0.02) (Figure 1). In dysbiosis situation, one luxR receptor gene from Bacteroides fragilis appeared to be over-expressed (p = 0.04) compared to non-dysbiotic patients (Figure 2). Conclusion Through this computational approach, AHL-synthesising bacteria have not been found. However, the expression of quorum sensing receptor genes appears to be modulated by IBD-associated gut dysbiosis. The role of LuxR receptors, especially in Bacteroides species, should be investigated to understand its impact on gut microbiota (Figure 3). Targeting LuxR receptors of bacterial quorum sensing might represent a new approach to modulate the gut microbiota in IBD.

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Callahan, Sean M., e Paul V. Dunlap. "LuxR- and Acyl-Homoserine-Lactone-Controlled Non-luxGenes Define a Quorum-Sensing Regulon in Vibrio fischeri". Journal of Bacteriology 182, n. 10 (15 maggio 2000): 2811–22. http://dx.doi.org/10.1128/jb.182.10.2811-2822.2000.

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ABSTRACT The luminescence (lux) operon (luxICDABEG) of the symbiotic bacterium Vibrio fischeri is regulated by the transcriptional activator LuxR and two acyl-homoserine lactone (acyl-HSL) autoinducers (the luxI-dependent 3-oxo-hexanoyl-HSL [3-oxo-C6-HSL] and the ainS-dependent octanoyl-HSL [C8-HSL]) in a population density-responsive manner called quorum sensing. To identify quorum-sensing-regulated (QSR) proteins different from those encoded by lux genes, we examined the protein patterns of V. fischeri quorum-sensing mutants defective in luxI, ainS, andluxR by two-dimensional polyacrylamide gel electrophoresis. Five non-Lux QSR proteins, QsrP, RibB, AcfA, QsrV, and QSR 7, were identified; their production occurred preferentially at high population density, required both LuxR and 3-oxo-C6-HSL, and was inhibited by C8-HSL at low population density. The genes encoding two of the QSR proteins were characterized: qsrP directs cells to synthesize an apparently novel periplasmic protein, andribB is a homolog of the Escherichia coli gene for 3,4-dihydroxy-2-butanone 4-phosphate synthase, a key enzyme for riboflavin synthesis. The qsrP and ribBpromoter regions each contained a sequence similar to thelux operon lux box, a 20-bp region of dyad symmetry necessary for LuxR/3-oxo-C6-HSL-dependent activation oflux operon transcription. V. fischeri qsrP andribB mutants exhibited no distinct phenotype in culture. However, a qsrP mutant, in competition with its parent strain, was less successful in colonizing Euprymna scolopes, the symbiotic host of V. fischeri. The newly identified QSR genes, together with the lux operon, define a LuxR/acyl-HSL-responsive quorum-sensing regulon in V. fischeri.

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Shadel, G. S., e T. O. Baldwin. "Positive autoregulation of the Vibrio fischeri luxR gene. LuxR and autoinducer activate cAMP-catabolite gene activator protein complex-independent and -dependent luxR transcription." Journal of Biological Chemistry 267, n. 11 (aprile 1992): 7696–702. http://dx.doi.org/10.1016/s0021-9258(18)42571-9.

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25

Zhang, Jun, Bing Liu, Dan Gu, Yuan Hao, Mo Chen, Yue Ma, Xiaohui Zhou, David Reverter, Yuanxing Zhang e Qiyao Wang. "Binding site profiles and N-terminal minor groove interactions of the master quorum-sensing regulator LuxR enable flexible control of gene activation and repression". Nucleic Acids Research 49, n. 6 (8 marzo 2021): 3274–93. http://dx.doi.org/10.1093/nar/gkab150.

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Abstract (sommario):

Abstract LuxR is a TetR family master quorum sensing (QS) regulator activating or repressing expression of hundreds of genes that control collective behaviors in Vibrios with underlying mechanism unknown. To illuminate how this regulator controls expression of various target genes, we applied ChIP-seq and DNase I-seq technologies. Vibrio alginolyticus LuxR controls expression of ∼280 genes that contain either symmetric palindrome (repDNA) or asymmetric (actDNA) binding motifs with different binding profiles. The median number of LuxR binding sites for activated genes are nearly double for that of repressed genes. Crystal structures of LuxR in complex with the respective repDNA and actDNA motifs revealed a new mode of LuxR DNA binding that involves contacts of its N-terminal extension to the minor groove. The N-terminal contacts mediated by Arginine-9 and Arginine-11 differ when LuxR binds to repDNA vs actDNA, leading to higher binding affinity at repressed targets. Moreover, modification of LuxR binding sites, binding profiles, and N-terminal extension have important consequences on QS-regulated phenotypes. These results facilitate fundamental understanding of the high flexibility of mechanisms of LuxR control of gene activation and repression in Vibrio QS, which may facilitate to design QS inhibiting chemicals that interfere with LuxR regulation to effectively control pathogens.

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Trott, Amy E., e Ann M. Stevens. "Amino Acid Residues in LuxR Critical for Its Mechanism of Transcriptional Activation during Quorum Sensing inVibrio fischeri". Journal of Bacteriology 183, n. 1 (1 gennaio 2001): 387–92. http://dx.doi.org/10.1128/jb.183.1.387-392.2001.

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ABSTRACT PCR-based site-directed mutagenesis has been used to generate 38 alanine-substitution mutations in the C-terminal 41 amino acid residues of LuxR. This region plays a critical role in the mechanism of LuxR-dependent transcriptional activation of the Vibrio fischeri lux operon during quorum sensing. The ability of the variant forms of LuxR to activate transcription of the lux operon was examined by using in vivo assays in recombinant Escherichia coli. Eight recombinant strains produced luciferase at levels less than 50% of that of a strain expressing wild-type LuxR. Western immunoblotting analysis verified that the altered forms of LuxR were expressed at levels equivalent to those of the wild type. An in vivo DNA binding-repression assay in recombinant E. coli was subsequently used to measure the ability of the variant forms of LuxR to bind to the lux box, the binding site of LuxR at thelux operon promoter. All eight LuxR variants found to affect cellular luciferase levels were unable to bind to thelux box. An additional 11 constructs that had no effect on cellular luciferase levels were also found to exhibit a defect in DNA binding. None of the alanine substitutions in LuxR affected activation of transcription of the lux operon without also affecting DNA binding. These results support the conclusion that the C-terminal 41 amino acids of LuxR are important for DNA recognition and binding of the lux box rather than positive control of the process of transcription initiation.

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Egland, Kristi A., e E. P. Greenberg. "Conversion of the Vibrio fischeriTranscriptional Activator, LuxR, to a Repressor". Journal of Bacteriology 182, n. 3 (1 febbraio 2000): 805–11. http://dx.doi.org/10.1128/jb.182.3.805-811.2000.

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Abstract (sommario):

ABSTRACT The Vibrio fischeri luminescence (lux) operon is regulated by a quorum-sensing system that involves the transcriptional activator (LuxR) and an acyl-homoserine lactone signal. Transcriptional activation requires the presence of a 20-base inverted repeat termed the lux box at a position centered 42.5 bases upstream of the transcriptional start of the lux operon. LuxR has proven difficult to study in vitro. A truncated form of LuxR has been purified, and together with ς70 RNA polymerase it can activate transcription of the lux operon. Both the truncated LuxR and RNA polymerase are required for binding tolux regulatory DNA in vitro. We have constructed an artificial lacZ promoter with the lux box positioned between and partially overlapping the consensus −35 and −10 hexamers of an RNA polymerase binding site. LuxR functioned as an acyl-homoserine lactone-dependent repressor at this promoter in recombinant Escherichia coli. Furthermore, multiplelux boxes on an independent replicon reduced the repressor activity of LuxR. Thus, it appears that LuxR can bind tolux boxes independently of RNA polymerase binding to the promoter region. A variety of LuxR mutant proteins were studied, and with one exception there was a correlation between function as a repressor of the artificial promoter and activation of a nativelux operon. The exception was the truncated protein that had been purified and studied in vitro. This protein functioned as an activator but not as a repressor in E. coli. The data indicate that the mutual dependence of purified, truncated LuxR and RNA polymerase on each other for binding to the lux promoter is a feature specific to the truncated LuxR and that full-length LuxR by itself can bind to lux box-containing DNA.

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Li, Shuangjia, Shijun Wu, Yixuan Ren, Qiu Meng, Jianhua Yin e Zhiliang Yu. "Characterization of differentiated autoregulation of LuxI/LuxR-type quorum sensing system in Pseudoalteromonas". Biochemical and Biophysical Research Communications 590 (gennaio 2022): 177–83. http://dx.doi.org/10.1016/j.bbrc.2021.12.107.

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Lau, Yin Yin, Kah Yan How, Wai-Fong Yin e Kok-Gan Chan. "Functional characterization of quorum sensing LuxR-type transcriptional regulator, EasR in Enterobacter asburiae strain L1". PeerJ 8 (21 ottobre 2020): e10068. http://dx.doi.org/10.7717/peerj.10068.

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Over the past decades, Enterobacter spp. have been identified as challenging and important pathogens. The emergence of multidrug-resistant Enterobacteria especially those that produce Klebsiella pneumoniae carbapenemase has been a very worrying health crisis. Although efforts have been made to unravel the complex mechanisms that contribute to the pathogenicity of different Enterobacter spp., there is very little information associated with AHL-type QS mechanism in Enterobacter spp. Signaling via N-acyl homoserine lactone (AHL) is the most common quorum sensing (QS) mechanism utilized by Proteobacteria. A typical AHL-based QS system involves two key players: a luxI gene homolog to synthesize AHLs and a luxR gene homolog, an AHL-dependent transcriptional regulator. These signaling molecules enable inter-species and intra-species interaction in response to external stimuli according to population density. In our recent study, we reported the genome of AHL-producing bacterium, Enterobacter asburiae strain L1. Whole genome sequencing and in silico analysis revealed the presence of a pair of luxI/R genes responsible for AHL-type QS, designated as easI/R, in strain L1. In a QS system, a LuxR transcriptional protein detects and responds to the concentration of a specific AHL controlling gene expression. In E. asburiae strain L1, EasR protein binds to its cognate AHLs, N-butanoyl homoserine lactone (C4-HSL) and N–hexanoyl homoserine lactone (C6-HSL), modulating the expression of targeted genes. In this current work, we have cloned the 693 bp luxR homolog of strain L1 for further characterization. The functionality and specificity of EasR protein in response to different AHL signaling molecules to activate gene transcription were tested and validated with β-galactosidase assays. Higher β-galactosidase activities were detected for cells harboring EasR, indicating EasR is a functional transcriptional regulator. This is the first report documenting the cloning and characterization of transcriptional regulator, luxR homolog of E. asburiae.

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Duerkop, Breck A., Ricky L. Ulrich e E. Peter Greenberg. "Octanoyl-Homoserine Lactone Is the Cognate Signal for Burkholderia mallei BmaR1-BmaI1 Quorum Sensing". Journal of Bacteriology 189, n. 14 (11 maggio 2007): 5034–40. http://dx.doi.org/10.1128/jb.00317-07.

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Abstract (sommario):

ABSTRACT Acyl-homoserine lactones (HSLs) serve as quorum-sensing signals for many Proteobacteria. Members of the LuxI family of signal generators catalyze the production of acyl-HSLs, which bind to a cognate receptor in the LuxR family of transcription factors. The obligate animal pathogen Burkholderia mallei produces several acyl-HSLs, and the B. mallei genome has four luxR and two luxI homologs, each of which has been established as a virulence factor. To begin to delineate the relevant acyl-HSL signals for B. mallei LuxR homologs, we analyzed the BmaR1-BmaI1 system. A comparison of acyl-HSL profiles from B. mallei ATCC 23344 and a B. mallei bmaI1 mutant indicates that octanoyl-HSL synthesis is BmaI1 dependent. Furthermore, octanoyl-HSL is the predominant acyl-HSL produced by BmaI1 in recombinant Escherichia coli. The synthesis of soluble BmaR1 in recombinant E. coli requires octanoyl-HSL or decanoyl-HSL. Insoluble aggregates of BmaR1 are produced in the presence of other acyl-HSLs and in the absence of acyl-HSLs. The bmaI1 promoter is activated by BmaR1 and octanoyl-HSL, and a 20-bp inverted repeat in the bmaI1 promoter is required for bmaI1 activation. Purified BmaR1 binds to this promoter region. These findings implicate octanoyl-HSL as the signal for BmaR1-BmaI1 quorum sensing and show that octanoyl-HSL and BmaR1 activate bmaI1 transcription.

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Chandler, Josephine R., Breck A. Duerkop, Aaron Hinz, T. Eoin West, Jake P. Herman, Mair E. A. Churchill, Shawn J. Skerrett e E. Peter Greenberg. "Mutational Analysis of Burkholderia thailandensis Quorum Sensing and Self-Aggregation". Journal of Bacteriology 191, n. 19 (31 luglio 2009): 5901–9. http://dx.doi.org/10.1128/jb.00591-09.

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Abstract (sommario):

ABSTRACT Acyl-homoserine lactone (acyl-HSL) quorum-sensing signaling is common to many Proteobacteria. Acyl-HSLs are synthesized by the LuxI family of synthases, and the signal response is mediated by members of the LuxR family of transcriptional regulators. Burkholderia thailandensis is a member of a closely related cluster of three species, including the animal pathogens Burkholderia mallei and Burkholderia pseudomallei. Members of this group have similar luxI and luxR homologs, and these genes contribute to B. pseudomallei and B. mallei virulence. B. thailandensis possesses three pairs of luxI-luxR homologs. One of these pairs, BtaI2-BtaR2, has been shown to produce and respond to 3OHC10-HSL and to control the synthesis of an antibiotic. By using a markerless-exhange method, we constructed an assortment of B. thailandensis quorum-sensing mutants, and we used these mutants to show that BtaI1 is responsible for C8-HSL production and BtaI3 is responsible for 3OHC8-HSL production. We also show that a strain incapable of acyl-HSL production is capable of growth on the same assortment of carbon and nitrogen sources as the wild type. Furthermore, this mutant shows no loss of virulence compared to the wild type in mice. However, the wild type self-aggregates in minimal medium, whereas the quorum-sensing mutant does not. The wild-type aggregation phenotype is recovered by addition of the BtaI1-R1 HSL signal C8-HSL. We propose that the key function of the BtaR1-BtaI1 quorum-sensing system is to cause cells to gather into aggregates once a sufficient population has been established.

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Joshi, Janak Raj, Netaly Khazanov, Amy Charkowski, Adi Faigenboim, Hanoch Senderowitz e Iris Yedidia. "Interkingdom Signaling Interference: The Effect of Plant-Derived Small Molecules on Quorum Sensing in Plant-Pathogenic Bacteria". Annual Review of Phytopathology 59, n. 1 (25 agosto 2021): 153–90. http://dx.doi.org/10.1146/annurev-phyto-020620-095740.

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Abstract (sommario):

In the battle between bacteria and plants, bacteria often use a population density–dependent regulatory system known as quorum sensing (QS) to coordinate virulence gene expression. In response, plants use innate and induced defense mechanisms that include low-molecular-weight compounds, some of which serve as antivirulence agents by interfering with the QS machinery. The best-characterized QS system is driven by the autoinducer N-acyl-homoserine lactone (AHL), which is produced by AHL synthases (LuxI homologs) and perceived by response regulators (LuxR homologs). Several plant compounds have been shown to directly inhibit LuxI or LuxR. Gaining atomic-level insight into their mode of action and how they interfere with QS enzymes supports the identification and design of novel QS inhibitors.Such information can be gained by combining experimental work with molecular modeling and docking simulations. The summary of these findings shows that plant-derived compounds act as interkingdom cues and that these allomones specifically target bacterial communication systems.

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Sayut, Daniel J., Yan Niu e Lianhong Sun. "Construction and Enhancement of a Minimal Genetic AND Logic Gate". Applied and Environmental Microbiology 75, n. 3 (5 dicembre 2008): 637–42. http://dx.doi.org/10.1128/aem.01684-08.

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ABSTRACT The ability of genetic networks to integrate multiple inputs in the generation of cellular responses is critical for the adaptation of cellular phenotype to distinct environments and of great interest in the construction of complex artificial circuits. To develop artificial genetic circuits that can integrate intercellular signaling molecules and commonly used inducing agents, we have constructed an artificial genetic AND gate based on the P luxI quorum-sensing promoter and the lac repressor. The hybrid promoter exhibited reduced basal and induced expression levels but increased expression capacity, generating clear logical responses that could be described using a simple mathematical model. The model also predicted that the AND gate's logic could be improved by altering the properties of the LuxR transcriptional activator and, in particular, by increasing its rate of transcriptional activation. Following these predictions, we were able to improve the AND gate's logic by ∼1.5-fold using a LuxR mutant library generated by directed evolution, providing the first example of the use of mutant transcriptional activators to improve the logic of a complex regulatory circuit. In addition, detailed characterizations of the AND gate's responses shed light on how LuxR, LacI, and RNA polymerase interact to activate gene expression.

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Finney, Angela H., Robert J. Blick, Katsuhiko Murakami, Akira Ishihama e Ann M. Stevens. "Role of the C-Terminal Domain of the Alpha Subunit of RNA Polymerase in LuxR-Dependent Transcriptional Activation of the lux Operon during Quorum Sensing". Journal of Bacteriology 184, n. 16 (15 agosto 2002): 4520–28. http://dx.doi.org/10.1128/jb.184.16.4520-4528.2002.

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Abstract (sommario):

ABSTRACT During quorum sensing in Vibrio fischeri, the luminescence, or lux, operon is regulated in a cell density-dependent manner by the activator LuxR in the presence of an acylated homoserine lactone autoinducer molecule [N-(3-oxohexanoyl) homoserine lactone]. LuxR, which binds to the lux operon promoter at a position centered at −42.5 relative to the transcription initiation site, is thought to function as an ambidextrous activator making multiple contacts with RNA polymerase (RNAP). The specific role of the α-subunit C-terminal domain (αCTD) of RNAP in LuxR-dependent transcriptional activation of the lux operon promoter has been investigated. The effects of 70 alanine substitution variants of the α subunit were determined in vivo by measuring the rate of transcription of the lux operon via luciferase assays in recombinant Escherichia coli. The mutant RNAPs from strains exhibiting at least twofold-increased or -decreased activity in comparison to the wild type were further examined by in vitro assays. Since full-length LuxR has not been purified, an autoinducer-independent N-terminally truncated form of LuxR, LuxRΔN, was used for in vitro studies. Single-round transcription assays were performed using reconstituted mutant RNAPs in the presence of LuxRΔN, and 14 alanine substitutions in the αCTD were identified as having negative effects on the rate of transcription from the lux operon promoter. Five of these 14 α variants were also involved in the mechanisms of both LuxR- and LuxRΔN-dependent activation in vivo. The positions of these residues lie roughly within the 265 and 287 determinants in α that have been identified through studies of the cyclic AMP receptor protein and its interactions with RNAP. This suggests a model where residues 262, 265, and 296 in α play roles in DNA recognition and residues 290 and 314 play roles in α-LuxR interactions at the lux operon promoter during quorum sensing.

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Zheng, Huiming, Zengtao Zhong, Xin Lai, Wen-Xin Chen, Shunpeng Li e Jun Zhu. "A LuxR/LuxI-Type Quorum-Sensing System in a Plant Bacterium, Mesorhizobium tianshanense, Controls Symbiotic Nodulation". Journal of Bacteriology 188, n. 5 (1 marzo 2006): 1943–49. http://dx.doi.org/10.1128/jb.188.5.1943-1949.2006.

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ABSTRACT The ability of rhizobia to symbiotically fix nitrogen from the atmosphere when forming nodules on their plant hosts requires various signal transduction pathways. LuxR-LuxI-type quorum-sensing systems have been shown to be one of the players in a number of rhizobium species. In this study, we found that Mesorhizobium tianshanense, a moderate-growth Rhizobium that forms nodules on a number of licorice plants, produces multiple N-acyl homoserine lactone (AHL)-like molecules. A simple screen for AHL synthase genes using an M. tianshanense genomic expression library in Escherichia coli, coupled with a sensitive AHL detector, uncovered a LuxI-type synthase, MrtI, and a LuxR-type regulator, MrtR, in M. tianshanense. Deletions of the mrtI or mrtR locus completely abolished AHL production in M. tianshanense. Using lacZ transcriptional fusions, we found that expression of the quorum-sensing regulators is autoinduced, as mrtI gene expression requires MrtR and cognate AHLs and mrtR expression is dependent on AHLs. Compared with the wild-type strains, quorum-sensing-deficient mutants showed a marked reduction in the efficiency of root hair adherence and, more importantly, were defective in nodule formation on their host plant, Glycyrrhiza uralensis. These data provide strong evidence that quorum sensing plays a critical role in the M. tianshanense symbiotic process.

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Qin, Nan, Sean M. Callahan, Paul V. Dunlap e Ann M. Stevens. "Analysis of LuxR Regulon Gene Expression during Quorum Sensing in Vibrio fischeri". Journal of Bacteriology 189, n. 11 (30 marzo 2007): 4127–34. http://dx.doi.org/10.1128/jb.01779-06.

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Abstract (sommario):

ABSTRACT The regulation of the lux operon (luxICDABEG) of Vibrio fischeri has been intensively studied as a model for quorum sensing in proteobacteria. Two-dimensional sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis previously identified several non-Lux proteins in V. fischeri MJ-100 whose expression was dependent on LuxR and 3-oxo-hexanoyl-l-homoserine lactone (3-oxo-C6-HSL). To determine if the LuxR-dependent regulation of the genes encoding these proteins was due to direct transcriptional control by LuxR and 3-oxo-C6-HSL or instead was due to indirect control via an unidentified regulatory element, promoters of interest were cloned into a lacZ reporter and tested for their LuxR and 3-oxo-C6-HSL dependence in recombinant Escherichia coli. The promoters for qsrP, acfA, and ribB were found to be directly activated via LuxR-3-oxo-C6-HSL. The sites of transcription initiation were established via primer extension analysis. Based on this information and the position of the lux box-binding site near position −40, all three promoters appear to have a class II-type promoter structure. In order to more fully characterize the LuxR regulon in V. fischeri MJ-100, real-time reverse transcription-PCR was used to study the temporal expression of qsrP, acfA, and ribB during the exponential and stationary phases of growth, and electrophoretic mobility shift assays were used to compare the binding affinities of LuxR to the promoters under investigation. Taken together, the results demonstrate that regulation of the production of QsrP, RibB, and AcfA is controlled directly by LuxR at the level of transcription, thereby establishing that there is a LuxR regulon in V. fischeri MJ-100 whose genes are coordinately expressed during mid-exponential growth.

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37

Egland, Kristi A., e E. P. Greenberg. "Quorum Sensing in Vibrio fischeri: Analysis of the LuxR DNA Binding Region by Alanine-Scanning Mutagenesis". Journal of Bacteriology 183, n. 1 (1 gennaio 2001): 382–86. http://dx.doi.org/10.1128/jb.183.1.382-386.2001.

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ABSTRACT LuxR is the transcriptional activator for quorum-sensing control of luminescence in Vibrio fischeri. A series of alanine-scanning mutants spanning a predicted helix-turn-helix region in the DNA binding domain of LuxR was constructed, and the activity of each of the LuxR mutant proteins in recombinant Escherichia coli was investigated. The region covered by the mutagenesis spanned residues 190 to 224. About half of the alanine-scanning mutants showed activities similar to that of the wild-type LuxR: at least two were positive-control mutants, four appeared to be defective in DNA binding, and several others were characterized as DNA binding affinity mutants. This analysis, taken together with information about other bacterial transcription factors, provides insights into amino acid residues in LuxR that are involved in DNA binding and transcriptional activation.

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38

Chatterjee, Jaidip, Carol M. Miyamoto e Edward A. Meighen. "Autoregulation of luxR: the Vibrio harveyi lux-operon activator functions as a repressor". Molecular Microbiology 20, n. 2 (aprile 1996): 415–25. http://dx.doi.org/10.1111/j.1365-2958.1996.tb02628.x.

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39

Patankar, Arati V., e Juan E. González. "Orphan LuxR regulators of quorum sensing". FEMS Microbiology Reviews 33, n. 4 (luglio 2009): 739–56. http://dx.doi.org/10.1111/j.1574-6976.2009.00163.x.

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40

Wu, Shijun, Shuangjia Li, Jianhua Yin e Zhiliang Yu. "Hfq and sRNA00002 positively regulate the LuxI/LuxR-type quorum sensing system in Pseudoalteromonas". Biochemical and Biophysical Research Communications 571 (settembre 2021): 1–7. http://dx.doi.org/10.1016/j.bbrc.2021.07.058.

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41

Fuqua, W. C., S. C. Winans e E. P. Greenberg. "Quorum sensing in bacteria: the LuxR-LuxI family of cell density-responsive transcriptional regulators." Journal of Bacteriology 176, n. 2 (1994): 269–75. http://dx.doi.org/10.1128/jb.176.2.269-275.1994.

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42

Hao, Youai, Stephen C. Winans, Bernard R. Glick e Trevor C. Charles. "Identification and characterization of new LuxR/LuxI-type quorum sensing systems from metagenomic libraries". Environmental Microbiology 12, n. 1 (gennaio 2010): 105–17. http://dx.doi.org/10.1111/j.1462-2920.2009.02049.x.

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43

Stevens, Ann M., Nobuyuki Fujita, Akira Ishihama e E. P. Greenberg. "Involvement of the RNA Polymerase α-Subunit C-Terminal Domain in LuxR-Dependent Activation of the Vibrio fischeri Luminescence Genes". Journal of Bacteriology 181, n. 15 (1 agosto 1999): 4704–7. http://dx.doi.org/10.1128/jb.181.15.4704-4707.1999.

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ABSTRACT LuxR is a ς70 RNA polymerase (RNAP)-dependent transcriptional activator that controls expression of the Vibrio fischeri lux operon in response to an acylhomoserine lactone-cell density signal. We have investigated whether the α-subunit C-terminal domain (αCTD) of RNAP is required for LuxR activity. A purified signal-independent, LuxR C-terminal domain-containing polypeptide (LuxRΔN) was used to study the activation of transcription from theluxI promoter in vitro. Initiation of luxoperon transcription was observed in the presence of LuxRΔN and wild-type RNAP but not in the presence of LuxRΔN and RNAPs with truncated αCTDs. We also studied the in vivo role of the RNAP αCTD in activation of lux transcription in Escherichia coli. This enabled a comparison of results obtained with full-length LuxR to those obtained with LuxRΔN. These in vivo studies indicated that both LuxR and LuxRΔN require the RNAP αCTD for activity. The results of DNase I protection studies showed that LuxRΔN-RNAP complexes can bind and protect the luxIpromoter, but with less efficacy when the αCTD is truncated in comparison to the wild type. Thus, both in vitro and in vivo experiments demonstrated that LuxR-dependent transcriptional activation of the lux operon involves the RNAP αCTD and suggest that αCTD-LuxR interactions may play a role in recruitment of RNAP to theluxI promoter.

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44

Costa, José M., e Joyce E. Loper. "Ecbl and EcbR: homologs of Luxl and LuxR affecting antibiotic and exoenzyme production byErwinia carotovorasubsp.betavasculorum". Canadian Journal of Microbiology 43, n. 12 (1 dicembre 1997): 1164–71. http://dx.doi.org/10.1139/m97-165.

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Erwinia carotovora subsp. betavasculorum Ecb168 causes vascular necrosis and root rot of sugar beet and produces an antibiotic(s) that is antagonistic against other Erwinia spp. EcbI−mutants of Ecb168, each containing a single transposon insertion in the ecbI gene (for Erwinia carotovora subsp. betavasculorum inducer), do not produce detectable levels of extracellular protease or antibiotic(s), and express less pectate lyase activity and virulence than the wild-type strain. A plasmid containing the cloned ecbI gene complemented the EcbI−mutants for these phenotypes. Protease production by EcbI−mutants grown on agar surfaces was restored by neighboring cells of Escherichia coli containing ecbI. Production of a diffusible N-acylhomoserine lactone autoinducer by wild-type Ecb168 was detected with indicator strains of E. coli and Agrobacterium tumefaciens. EcbI−mutant strains did not produce an autoinducer detected by the indicator strains. Antibiotic production by EcbI−mutants was restored by cell-free culture supernatants of Ecb168 or E. coli containing a cloned ecbI gene. The predicted amino acid sequence of EcbI is similar to those of CarI, ExpI, and HslI, three LuxI homologs required for production of a diffusible N-acylhomoserine lactone autoinducer in Erwinia carotovora subsp. carotovora. A luxR homolog, termed ecbR (for Erwinia carotovora subsp. betavasculorum regulator), is convergently transcribed and overlaps with ecbI by 17 bp at their 3′ ends. These results are consistent with the hypothesis that a quorum-sensing system related to the prototypic luxI–luxR gene pair controls antibiotic and exoenzyme production in Erwinia carotovora subsp. betavasculorum.Key words: quorum sensing, β-lactam, gene regulation.

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45

Manukhov, Ilya V., Ol'ga E. Melkina, Ignatii I. Goryanin, Ancha V. Baranova e Gennadii B. Zavilgelsky. "The N-Terminal Domain of Aliivibrio fischeri LuxR Is a Target of the GroEL Chaperonin". Journal of Bacteriology 192, n. 20 (20 agosto 2010): 5549–51. http://dx.doi.org/10.1128/jb.00754-10.

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ABSTRACT Here we show that the C-terminal domain of LuxR activates the transcription of Aliivibrio fischeri luxICDABEG in Escherichia coli SKB178 gro + and E. coli OFB1111 groEL673 strains to the same level. Using affinity chromatography, we showed that GroEL binds to the N-terminal domain of LuxR, pointing to a GroEL/GroES requirement for the folding of the N-terminal domain of LuxR.

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46

Kviatkovski, I., T. Yarnitzky, S. Shushan, O. Schwartz-Harari, R. Nir-Paz e Y. Helman. "A bacterial biosensor encoding a genetically modified LuxR receptor exhibits improved detection of Pseudomonas aeruginosa's biomarker molecule 2-aminoacetophenone". Chemical Communications 54, n. 66 (2018): 9218–21. http://dx.doi.org/10.1039/c8cc03540g.

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47

Malott, Rebecca J., Eoin P. O'Grady, Jessica Toller, Silja Inhülsen, Leo Eberl e Pamela A. Sokol. "A Burkholderia cenocepacia Orphan LuxR Homolog Is Involved in Quorum-Sensing Regulation". Journal of Bacteriology 191, n. 8 (6 febbraio 2009): 2447–60. http://dx.doi.org/10.1128/jb.01746-08.

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ABSTRACT Burkholderia cenocepacia utilizes quorum sensing to control gene expression, including the expression of genes involved in virulence. In addition to CepR and CciR, a third LuxR homolog, CepR2, was found to regulate gene expression and virulence factor production. All B. cenocepacia strains examined contained this orphan LuxR homolog, which was not associated with an adjacent N-acyl-homoserine lactone synthase gene. Expression of cepR2 was negatively autoregulated and was negatively regulated by CciR in strain K56-2. Microarray analysis and quantitative reverse transcription-PCR determined that CepR2 did not influence expression of cepIR or cciIR. However, in strain K56-2, CepR2 negatively regulated expression of several known quorum-sensing-controlled genes, including genes encoding zinc metalloproteases. CepR2 exerted positive and negative regulation on genes on three chromosomes, including strong negative regulation of a gene cluster located adjacent to cepR2. In strain H111, which lacks the CciIR quorum-sensing system, CepR2 positively regulated pyochelin production by controlling transcription of one of the operons required for the biosynthesis of the siderophore in an N-acyl-homoserine lactone-independent manner. CepR2 activation of a luxI promoter was demonstrated in a heterologous Escherichia coli host, providing further evidence that CepR2 can function in the absence of signaling molecules. This study demonstrates that the orphan LuxR homolog CepR2 contributes to the quorum-sensing regulatory network in two distinct strains of B. cenocepacia.

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48

Freeman, Jeremy A., e Bonnie L. Bassler. "Sequence and Function of LuxU: a Two-Component Phosphorelay Protein That Regulates Quorum Sensing inVibrio harveyi". Journal of Bacteriology 181, n. 3 (1 febbraio 1999): 899–906. http://dx.doi.org/10.1128/jb.181.3.899-906.1999.

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ABSTRACT Vibrio harveyi regulates the expression of bioluminescence (lux) in response to cell density, a phenomenon known as quorum sensing. In V. harveyi, two independent quorum-sensing systems exist, and each produces, detects, and responds to a specific cell density-dependent autoinducer signal. The autoinducers are recognized by two-component hybrid sensor kinases called LuxN and LuxQ, and sensory information from both systems is transduced by a phosphorelay mechanism to the response regulator protein LuxO. Genetic evidence suggests that LuxO-phosphate negatively regulates the expression of luminescence at low cell density in the absence of autoinducers. At high cell density, interaction of the sensors with their cognate autoinducers results in dephosphorylation and inactivation of the LuxO repressor. In the present report, we show that LuxN and LuxQ channel sensory information to LuxO via a newly identified phosphorelay protein that we have named LuxU. LuxU shows sequence similarity to other described phosphorelay proteins, including BvgS, ArcB, and Ypd1. A critical His residue (His 58) of LuxU is required for phosphorelay function.

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49

Fuqua, Clay, Stephen C. Winans e E. Peter Greenberg. "CENSUS AND CONSENSUS IN BACTERIAL ECOSYSTEMS: The LuxR-LuxI Family of Quorum-Sensing Transcriptional Regulators". Annual Review of Microbiology 50, n. 1 (ottobre 1996): 727–51. http://dx.doi.org/10.1146/annurev.micro.50.1.727.

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50

Miyamoto, Carol M., Paul V. Dunlap, Edward G. Ruby e Edward A. Meighen. "LuxO controls luxR expression in Vibrio harveyi: evidence for a common regulatory mechanism in Vibrio". Molecular Microbiology 48, n. 2 (4 aprile 2003): 537–48. http://dx.doi.org/10.1046/j.1365-2958.2003.03453.x.

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