954 resultados para Quorum quenching


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The implications of Quorum Sensing in the establishment of a successful symbiosis of Rhizobium leguminosarum bv. viciae (Rlv) with legume plants are discussed in this work. In order to analyze the significance and regulation of the production of AHL signal molecules, mutants deficient in each of the two QS systems present in Rlv UPM791 were constructed. A detailed analysis of the effect of these mutations on growth, AHL production, biofilm formation and symbiosis with pea, vetch and lentil plants has been carried out.

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Rhizobium leguminosarum bv viciae (Rlv) es una alfa-proteobacteria capaz de establecer una simbiosis diazotrófica con distintas leguminosas. Uno de los factores implicados en el establecimiento de la simbiosis es el sistema de comunicación intercelular conocido como Quorum Sensing (QS). Mediante este sistema, las bacterias actúan de manera coordinada en respuesta a cambios en la densidad de población a través de la producción y detección de señales extracelulares. El genoma de Rlv UPM791 contiene dos sistemas tipo luxRI mediados por señales de tipo N-acyl-homoserina lactonas (AHLs): el sistema rhiRI, codificado en el plásmido simbiótico, produce C6-HSL, C7-HSL y C8-HSL; y el sistema cinRI, localizado en el cromosoma, produce 3-OH-C14:1-HSL. Con el fin de analizar el significado y la regulación de los sistemas de QS en esta bacteria endosimbiótica se generaron mutantes defectivos en cada uno de los sistemas de QS, y se llevó a cabo un análisis detallado sobre la producción de AHLs y la simbiosis con plantas de guisante, veza y lenteja. El sistema rhiRI se necesita para un comportamiento simbiótico normal, dado que la mutación en rhiI reduce considerablemente la eficiencia simbiótica. rhiR es esencial para la fijación de nitrógeno en ausencia del plásmido pUPM791d. Asimismo, mutaciones en el sistema cinRIS mostraron también un importante efecto en simbiosis. El mutante ?cinRIS no produce la señal 3-OH-C14:1-HSL, y da lugar a nódulos blancos e inefectivos, carentes de bacteroides. El mutante ?cinI, incapaz de producir AHLs, no forma nódulos en ninguna de las leguminosas utilizadas. El análisis genético reveló que dicha mutación origina la inestabilización del plásmido simbiótico por un mecanismo dependiente de cinI que no ha sido aclarado. Los resultados obtenidos sugieren un papel relevante de los sistemas de Quorum Sensing de Rlv UPM791 en los primeros estadíos de la simbiosis, e indican la existencia de un modelo de regulación dependiente de QS significativamente distinto a los que se han descrito previamente en otras cepas de R. leguminosarum.

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Copia digital: Biblioteca valenciana, 2010

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Classical quorum-sensing (autoinduction) regulation, as exemplified by the lux system of Vibrio fischeri, requires N-acyl homoserine lactone (AHL) signals to stimulate cognate transcriptional activators for the cell density-dependent expression of specific target gene systems. For Pantoea stewartii subsp. stewartii, a bacterial pathogen of sweet corn and maize, the extracellular polysaccharide (EPS) stewartan is a major virulence factor, and its production is controlled by quorum sensing in a population density-dependent manner. Two genes, esaI and esaR, encode essential regulatory proteins for quorum sensing. EsaI is the AHL signal synthase, and EsaR is the cognate gene regulator. esaI, ΔesaR, and ΔesaI-esaR mutations were constructed to establish the regulatory role of EsaR. We report here that strains containing an esaR mutation produce high levels of EPS independently of cell density and in the absence of the AHL signal. Our data indicate that quorum-sensing regulation in P. s. subsp. stewartii, in contrast to most other described systems, uses EsaR to repress EPS synthesis at low cell density, and that derepression requires micromolar amounts of AHL. In addition, derepressed esaR strains, which synthesize EPS constitutively at low cell densities, were significantly less virulent than the wild-type parent. This finding suggests that quorum sensing in P. s. subsp. stewartii may be a mechanism to delay the expression of EPS during the early stages of infection so that it does not interfere with other mechanisms of pathogenesis.

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Escherichia coli and Salmonella typhimurium strains grown in Luria–Bertani medium containing glucose secrete a small soluble heat labile organic molecule that is involved in intercellular communication. The factor is not produced when the strains are grown in Luria–Bertani medium in the absence of glucose. Maximal secretion of the substance occurs in midexponential phase, and the extracellular activity is degraded as the glucose is depleted from the medium or by the onset of stationary phase. Destruction of the signaling molecule in stationary phase indicates that, in contrast to other quorum-sensing systems, quorum sensing in E. coli and S. typhimurium is critical for regulating behavior in the prestationary phase of growth. Our results further suggest that the signaling factor produced by E. coli and S. typhimurium is used to communicate both the cell density and the metabolic potential of the environment. Several laboratory and clinical strains of E. coli and S. typhimurium were screened for production of the signaling molecule, and most strains make it under conditions similar to those shown here for E. coli AB1157 and S. typhimurium LT2. However, we also show that E. coli strain DH5α does not make the soluble factor, indicating that this highly domesticated strain has lost the gene(s) or biosynthetic machinery necessary to produce the signaling substance. Implications for the involvement of quorum sensing in pathogenesis are discussed.

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The fluorescence of a polyanionic conjugated polymer can be quenched by extremely low concentrations of cationic electron acceptors in aqueous solutions. We report a greater than million-fold amplification of the sensitivity to fluorescence quenching compared with corresponding “molecular excited states.” Using a combination of steady-state and ultrafast spectroscopy, we have established that the dramatic quenching results from weak complex formation [polymer(−)/quencher(+)], followed by ultrafast electron transfer from excitations on the entire polymer chain to the quencher, with a time constant of 650 fs. Because of the weak complex formation, the quenching can be selectively reversed by using a quencher-recognition diad. We have constructed such a diad and demonstrate that the fluorescence is fully recovered on binding between the recognition site and a specific analyte protein. In both solutions and thin films, this reversible fluorescence quenching provides the basis for a new class of highly sensitive biological and chemical sensors.

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Collectively, the xanthophyll class of carotenoids perform a variety of critical roles in light harvesting antenna assembly and function. The xanthophyll composition of higher plant photosystems (lutein, violaxanthin, and neoxanthin) is remarkably conserved, suggesting important functional roles for each. We have taken a molecular genetic approach in Arabidopsis toward defining the respective roles of individual xanthophylls in vivo by using a series of mutant lines that selectively eliminate and substitute a range of xanthophylls. The mutations, lut1 and lut2 (lut = lutein deficient), disrupt lutein biosynthesis. In lut2, lutein is replaced mainly by a stoichiometric increase in violaxanthin and antheraxanthin. A third mutant, aba1, accumulates normal levels of lutein and substitutes zeaxanthin for violaxanthin and neoxanthin. The lut2aba1 double mutant completely lacks lutein, violaxanthin, and neoxanthin and instead accumulates zeaxanthin. All mutants were viable in soil and had chlorophyll a/b ratios ranging from 2.9 to 3.5 and near wild-type rates of photosynthesis. However, mutants accumulating zeaxanthin exhibited a delayed greening virescent phenotype, which was most severe and often lethal when zeaxanthin was the only xanthophyll present. Chlorophyll fluorescence quenching kinetics indicated that both zeaxanthin and lutein contribute to nonphotochemical quenching; specifically, lutein contributes, directly or indirectly, to the rapid rise of nonphotochemical quenching. The results suggest that the normal complement of xanthophylls, while not essential, is required for optimal assembly and function of the light harvesting antenna in higher plants.

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Bacteria communicate with each other to coordinate expression of specific genes in a cell density-dependent fashion, a phenomenon called quorum sensing and response. Although we know that quorum sensing via acyl-homoserine lactone (HSL) signals controls expression of several virulence genes in the human pathogen Pseudomonas aeruginosa, the number and types of genes controlled by quorum sensing have not been studied systematically. We have constructed a library of random insertions in the chromosome of a P. aeruginosa acyl-HSL synthesis mutant by using a transposon containing a promoterless lacZ. This library was screened for acyl-HSL induction of lacZ. Thirty-nine quorum sensing-regulated genes were identified. The genes were organized into classes depending on the pattern of regulation. About half of the genes appear to be in seven operons, some seem organized in large patches on the genome. Many of the quorum sensing-regulated genes code for putative virulence factors or production of secondary metabolites. Many of the genes identified showed a high level of induction by acyl-HSL signaling.

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Enterohemorrhagic Escherichia coli O157:H7 and enteropathogenic E. coli cause a characteristic histopathology in intestinal cells known as attaching and effacing. The attaching and effacing lesion is encoded by the Locus of Enterocyte Effacement (LEE) pathogenicity island, which encodes a type III secretion system, the intimin intestinal colonization factor, and the translocated intimin receptor protein that is translocated from the bacterium to the host epithelial cells. Using lacZ reporter gene fusions, we show that expression of the LEE operons encoding the type III secretion system, translocated intimin receptor, and intimin is regulated by quorum sensing in both enterohemorrhagic E. coli and enteropathogenic E. coli. The luxS gene recently shown to be responsible for production of autoinducer in the Vibrio harveyi and E. coli quorum-sensing systems is responsible for regulation of the LEE operons, as shown by the mutation and complementation of the luxS gene. Regulation of intestinal colonization factors by quorum sensing could play an important role in the pathogenesis of disease caused by these organisms. These results suggest that intestinal colonization by E. coli O157:H7, which has an unusually low infectious dose, could be induced by quorum sensing of signals produced by nonpathogenic E. coli of the normal intestinal flora.