Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P51532 (transcriptional activator)
 6,546 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

A series of structural analogs of the Pseudomonas aeruginosa autoinducer [PAI, N-3-oxo-dodecanoyl homoserine lactone] were obtained and tested for their ability to act as autoinducers in stimulating the expression of the gene for elastase (lasB) by measuring beta-galactosidase production from a lasB-lacZ gene fusion in the presence of the transcriptional activator LasR. The data suggest that the length of the acyl side chain of the autoinducer molecule is the most critical factor for activity. Replacement of the ring O by S in the homoserine lactone moiety can be tolerated. Tritium-labelled PAI ([3H]PAI) was synthesized and used to demonstrate the association of [3H]PAI with cells overexpressing LasR. The PAI analogs were also tested for their ability to compete with [3H]PAI for binding of LasR. Results from the competition assays suggest that once again the length of the acyl side chain appears to be crucial for antagonist activity. The presence of the 3-oxo moiety also plays a significant role in binding since analogs which lacked this moiety were much less effective in blocking binding of [3H]PAI. All analogs demonstrating competition with PAI in binding to LasR also exhibited the ability to activate lasB expression, suggesting that they are functional analogs of PAI.
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PMID:Functional analysis of the Pseudomonas aeruginosa autoinducer PAI. 883 Jun 97

The importance of accurate demographic information is reflected in the United States Constitution, Article 1, which provides for a decennial census of this country's human population. Bacteria also conduct a census of their population and do so more frequently, more efficiently, and as far we know, with little if any of the political contentiousness caused by human demographers. Many examples have been found of particular bacterial genes, operons, or regulons that are expressed preferentially at high cell densities. Many of these are regulated by proteins related to the LuxR and LuxI proteins of Vibrio fischeri, and by a diffusible pheromone called an autoinducer. LuxR and LuxI and their cognate autoinducer (3-oxohexanoyl homoserine lactone, designated VAI-1) provide an important model to describe the functions of this family of proteins. LuxR is a VAI-1 receptor and a VAI-1-dependent transcriptional activator, and LuxI directs the synthesis of VAI-1. VAI-1 diffuses across the bacterial envelope, and intracellular concentrations of it are therefore strongly increased by nearby VAI-1-producing bacteria. Similar systems regulate pathogenesis factors in Pseudomonas aeruginosa and Erwinia spp., as well as T1 plasmid conjugal transfer in Agrobacterium tumefaciens, and many other genes in numerous genera of gram-negative bacteria. Genetic analyses of these systems have revealed a high degree of functional conservation, while also uncovering features that are unique to each.
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PMID:Census and consensus in bacterial ecosystems: the LuxR-LuxI family of quorum-sensing transcriptional regulators. 890 97

Two regions of the nopaline-type Ti plasmid pTiC58 are important for conjugal transfer of this element to recipient bacteria. These two regions were cloned into two independent replicons to produce a binary transfer system. For one region, oriT/tra, we constructed two derivatives, pFRtra and pDCtra-5. Each contains the oriT site and the two flanking, divergently transcribed tra operons that encode the DNA processing functions associated with the relaxosome. These two plasmids also carry traR, which encodes the transcriptional activator necessary for expression of transfer genes. The two plasmids differ by the amounts of traB sequence or sequence downstream of traG present in the construct. The second replicon, pPLE2, carries the traI/trb region. The traI gene confers production of the Agrobacterium tumefaciens N-acyl homoserine lactone autoinducer, while the remaining genes in the trb operon encode components of the mating bridge. Donors harboring the two plasmids mobilized the transfer of the plasmid carrying the oriT/tra region to an A. tumefaciens recipient at frequencies similar to that at which the intact Ti plasmid transferred. Plasmid pFRtra, which encodes most of traB, was mobilized at a frequency almost 10-fold higher than was pDCtra-5, which lacks most of the gene. A. tumefaciens donors also mobilized pFRtra to Escherichia coli and Pseudomonas fluorescens recipients at frequencies similar to those observed with A. tumefaciens recipients. Rhizobium meliloti harboring the binary system also transferred the oriT/tra component to these recipients. However, E. coli or P. fluorescens donors harboring the binary system did not transfer pFRtra to any of the recipients. Furthermore, while the A. tumefaciens and R. meliloti donors produced high levels of the autoinducer, the P. fluorescens and E. coli donors produced only trace amounts of this signal molecule. These results indicate that the tra system of pTiC58 is fully contained within the characterized tra and trb regions of the Ti plasmid, that conjugation does not require functions encoded by the vir system for maximal activity, and that while the Ti plasmid tra system recognizes diverse gram-negative bacteria as recipients, of the hosts tested, it functions only in members of the family Rhizobiaceae.
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PMID:Ti plasmid conjugation is independent of vir: reconstitution of the tra functions from pTiC58 as a binary system. 902 14

The global activator GacA, a highly conserved response regulator in Gram-negative bacteria, is required for the production of exoenzymes and secondary metabolites in Pseudomonas spp. The gacA gene of Pseudomonas aeruginosa PAO1 was isolated and its role in cell-density-dependent gene expression was characterized. Mutational inactivation of gacA resulted in delayed and reduced formation of the cell-density signal N-butyryl-L-homoserine lactone (BHL), of the cognate transcriptional activator RhIR (VsmR), and of the transcriptional activator LasR, which is known to positively regulate RhIR expression. Amplification of gacA on a multicopy plasmid caused precocious and enhanced production of BHL, RhIR and LasR. In parallel, the gacA gene dosage markedly influenced the BHL/RhIR-dependent formation of the cytotoxic compounds pyocyanin and cyanide and the exoenzyme lipase. However, the concentrations of another known cell-density signal of P. aeruginosa, N-oxododecanoyl-L-homoserine lactone, did not always match BHL concentrations. A model accounting for these observations places GacA function upstream of LasR and RhIR in the complex, cell-density-dependent signal-transduction pathway regulating several exoproducts and virulence factors of P. aeruginosa via BHL.
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PMID:The global activator GacA of Pseudomonas aeruginosa PAO positively controls the production of the autoinducer N-butyryl-homoserine lactone and the formation of the virulence factors pyocyanin, cyanide, and lipase. 915 18

Two quorum-sensing systems (las and rhl) regulate virulence gene expression in Pseudomonas aeruginosa. The las system consists of a transcriptional activator, LasR, and LasI, which directs the synthesis of the autoinducer N-(3-oxododecanoyl) homoserine lactone (PAI-1). Induction of lasB (encoding elastase) and other virulence genes requires LasR and PAI-1. The rhl system consists of a putative transcriptional activator, RhlR, and RhlI, which directs the synthesis of N-butyryl homoserine lactone (PAI-2). Rhamnolipid production in P. aeruginosa has been reported to require both the rhl system and rhlAB (encoding a rhamnosyltransferase). Here we report the generation of a delta lasI mutant and both delta lasI delta rhlI and delta lasR rhlR::Tn501 double mutants of strain PAO1. Rhamnolipid production and elastolysis were reduced in the delta lasI single mutant and abolished in the double-mutant strains. rhlAB mRNA was not detected in these strains at mid-logarithmic phase but was abundant in the parental strain. Further RNA analysis of the wild-type strain revealed that rhlAB is organized as an operon. The rhlAB transcriptional start was mapped, and putative sigma 54 and sigma 70 promoters were identified upstream. To define components required for rhlAB expression, we developed a bioassay in Escherichia coli and demonstrated that PAI-2 and RhlR are required and sufficient for expression of rhlA. To characterize the putative interaction between PAI-2 and RhlR, we demonstrated that [3H]PAI-2 binds to E. coli cells expressing RhlR and not to those expressing LasR. Finally, the specificity of the las and rhl systems was examined in E. coli bioassays. The las system was capable of mildly activating rhlA, and similarly, the rhl system partly activated lasB. However; these effects were much less than the activation of rhlA by the rhl system and lasB by the las system. The results presented here further characterize the roles of the rhl and las quorum-sensing systems in virulence gene expression.
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PMID:Roles of Pseudomonas aeruginosa las and rhl quorum-sensing systems in control of elastase and rhamnolipid biosynthesis genes. 929 32

Quorum sensing relies upon the interaction of a diffusible signal molecule with a transcriptional activator protein to couple gene expression with cell population density. In Gram-negative bacteria, such signal molecules are usually N-acylhomoserine lactones (AHLs) which differ in the structure of their N-acyl side chains. Chromobacterium violaceum, a Gram-negative bacterium commonly found in soil and water, produces the characteristic purple pigment violacein. Previously the authors described a violacein-negative, mini-Tn5 mutant of C. violaceum (CV026) in which pigment production can be restored by incubation with supernatants from the wild-type strain. To develop this mutant as a general biosensor for AHLs, the natural C. violaceum AHL molecule was first chemically characterized. By using solvent extraction, HPLC and mass spectrometry, a single AHL, N-hexanoyl-L-homoserine lactone (HHL), was identified in wild-type C. violaceum culture supernatants which was absent from CV026. Since the production of violacein constitutes a simple assay for the detection of AHLs, we explored the ability of CV026 to respond to a series of synthetic AHL and N-acylhomocysteine thiolactone (AHT) analogues. In CV026, violacein is inducible by all the AHL and AHT compounds evaluated with N-acyl side chains from C4 to C8 in length, with varying degrees of sensitivity. Although AHL compounds with N-acyl side chains from C10 to C14 are unable to induce violacein production, if an activating AHL (e.g. HHL) is incorporated into the agar, these long-chain AHLs can be detected by their ability to inhibit violacein production. The versatility of CV026 in facilitating detection of AHL mixtures extracted from culture supernatants and separated by thin-layer chromatography is also demonstrated. These simple bioassays employing CV026 thus greatly extend the ability to detect a wide spectrum of AHL signal molecules.
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PMID:Quorum sensing and Chromobacterium violaceum: exploitation of violacein production and inhibition for the detection of N-acylhomoserine lactones. 942 96

Conjugal transfer of Agrobacterium tumefaciens Ti plasmids is regulated by two hierarchical signalling systems. Transfer is dependent on a subset of opines produced by the plant tumours induced by the bacterium. Induction also requires an acyl-homoserine lactone signal, called AAI, that is produced by the bacteria themselves. AAI is the co-inducer for TraR, the transcriptional activator required for expression of the tra regulon. Octopine induces conjugation of the octopine-mannityl opine-type Ti plasmids by regulating the expression of traR via OccR, the octopine-dependent activator of the opine regulon. We have discovered a second traR-like gene, trlR, on the octopine-mannityl opine-type Ti plasmids pTi15955 and pTiR10. This gene is located in an operon coding for a mannopine transport system and is expressed as part of the mannityl opine regulon. Sequence analysis indicated that trlR is a frameshift allele of traR, and the resulting protein lacks the carboxy-terminal domain thought to constitute the DNA-binding region of TraR. Expression of trlR inhibited octopine-induced conjugation of pTi15955 and pTiR10 by suppressing the TraR-mediated transcription of the tra and trb operons. Although TrlR had no effect on the expression of traR, TraR activated the expression of trlR. Southern hybridizations indicated that several other Ti and opine-catabolic plasmids contain more than one copy of genes homologous to traR. We propose that trlR is a dominant negative allele of traR and that TrlR inhibits conjugation by forming inactive heteromultimers with TraR.
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PMID:Octopine-type Ti plasmids code for a mannopine-inducible dominant-negative allele of traR, the quorum-sensing activator that regulates Ti plasmid conjugal transfer. 948 84

Few strains of Erwinia carotovora subsp. carotovora (Ecc) make carbapenem antibiotics. Strain GS101 makes the basic carbapenem molecule, 1-carbapen-2-em-3-carboxylic acid (Car). The production of this antibiotic has been shown to be cell density dependent, requiring the accumulation of the small diffusible molecule N-(3-oxohexanoyl)-L-homoserine lactone (OHHL) in the growth medium. When the concentration of this inducer rises above a threshold level, OHHL is proposed to interact with the transcriptional activator of the carbapenem cluster (CarR) and induce carbapenem biosynthesis. The introduction of the GS101 carR gene into an Ecc strain (SCRI 193) which is naturally carbapenem-negative resulted in the production of Car. This suggested that strain SCRI 193 contained functional cryptic carbapenem biosynthetic genes, but lacked a functional carR homologue. The distribution of trans-activatable antibiotic genes was assayed in Erwinia strains from a culture collection and was found to be common in a large proportion of Ecc strains. Significantly, amongst the Ecc strains identified, a larger proportion contained trans-activatable cryptic genes than produced antibiotics constitutively. Southern hybridization of the chromosomal DNA of cryptic Ecc strains confirmed the presence of both the car biosynthetic cluster and the regulatory genes. Identification of homologues of the transcriptional activator carR suggests that the cause of the silencing of the carbapenem biosynthetic cluster in these strains is not the deletion of carR. In an attempt to identify the cause of the silencing in the Ecc strain SCRI 193 the carR homologue from this strain was cloned and sequenced. The SCRI 193 CarR homologue was 94% identical to the GS101 CarR and contained 14 amino acid substitutions. Both homologues could be expressed from their native promoters and ribosome-binding sites using an in vitro prokaryotic transcription and translation assay, and when the SCRI 193 carR homologue was cloned in multicopy plasmids and reintroduced into SCRI 193, antibiotic production was observed. This suggested that the mutation causing the silencing of the biosynthetic cluster in SCRI 193 was leaky and the cryptic Car phenotype could be suppressed by multiple copies of the apparently mutant transcriptional activator.
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PMID:Cryptic carbapenem antibiotic production genes are widespread in Erwinia carotovora: facile trans activation by the carR transcriptional regulator. 963 20

The TraR and TraI proteins of Agrobacterium tumefaciens mediate cell-density-dependent expression of the Ti plasmid tra regulon. TraI synthesizes the autoinducer pheromone N-(3-oxooctanoyl)-L-homoserine lactone (3-oxo-C8-HSL), while TraR is an 3-oxo-C8-HSL-responsive transcriptional activator. We have compared the abilities of 3-oxo-C8-HSL and 32 related compounds to activate expression of a TraR-regulated promoter. In a strain that expresses wild-type levels of TraR, only 3-oxo-C8-HSL was strongly stimulatory, four compounds were detectably active only at high concentrations, and the remaining 28 compounds were inactive. Furthermore, many of these compounds were potent antagonists. In contrast, almost all of these compounds were stimulatory in a congenic strain that overexpresses TraR and no compound was a potent antagonist. We propose a model in which autoinducers enhance the affinity of TraR either for other TraR monomers or for DNA binding sites and that overexpression of TraR potentiates this interaction by mass action. Wild-type A. tumefaciens released a rather broad spectrum of autoinducers, including several that antagonize induction of a wild-type strain. However, under all conditions tested, 3-oxo-C8-HSL was more abundant than any other analog, indicating that other released autoinducers do not interfere with tra gene induction. We conclude that (i) in wild-type strains, only 3-oxo-C8-HSL significantly stimulates tra gene expression, while many autoinducer analogs are potent antagonists; (ii) TraR overexpression increases agonistic activity of autoinducer analogs, allowing sensitive biodetection of many autoinducers; and (iii) autoinducer stimulatory activity is potentiated by TraR overproduction, suggesting that autoinducers may shift an equilibrium between TraR monomers and dimers or oligomers. When autoinducer specificities of other quorum-sensing proteins are tested, care should be taken not to overexpress those proteins.
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PMID:Analogs of the autoinducer 3-oxooctanoyl-homoserine lactone strongly inhibit activity of the TraR protein of Agrobacterium tumefaciens. 976 71

The recent discovery that the fish pathogen Vibrio salmonicida is closely related to the luminous bacteria Vibrio fischeri and Vibrio logei suggested that V. salmonicida might also be capable of bioluminescence. Interestingly, cells of V. salmonicida were found to produce light in culture, but only when exposed to either an aliphatic aldehyde and/or the major V. fischeri autoinducer N-(3-oxo-hexanoyl)-L-homoserine lactone, a transcriptional activator of the luminescence (lux) genes. An extract of spent medium of V. salmonicida that should contain any V. salmonicida acyl-homoserine lactone autoinducer, when added to V. fischeri cells, led to an induction of their luminescence. These results show that V. salmonicida is a newly recognized luminous bacterial species that apparently both produces an autoinducer activity and responds to exogenous V. fischeri autoinducer.
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PMID:Cryptic luminescence in the cold-water fish pathogen Vibrio salmonicida. 1020 Oct 98


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