3 quorum sensing signals add in microarray comparison in Yersinia pestis at 30°C
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ABSTRACT: The quorum-sensing system has been linked to diverse phenotypes and regulatory changes in pathogenic bacteria. In strain CO92, the AI-2 signal is produced in a luxS-dependent manner, reaching maximal levels of 2.5 μM in late logarithmic growth, and both wild type and pigmentation mutant strains made equivalent levels of AI-2. Yersinia pestis CO92 possesses a chromosomal lsr locus encoding factors involved in the binding and import of AI-2, and confirming this assignment, an lsr deletion increased extracellular pools of AI-2. To assess the functional role of 3 quorum sensing singlas in Y. pestis, microarray study was conducted comparing the 3 quorum sensing signals added in to control at 30°C to mimic the flea gut. Here, the ÎPgm (pigmentation-negative) R88 strain was used. The three signals are one AI-2 and two AHLs (N-(3-Oxooctanoyl)-L-homoserine lactone and N-Hexanoyl-DL-homoserine lactone).The control consisted of cells grown and treated under the same conditions without added signals. Six independent RNA samples from 3 signals add in cultures were paired with six independent RNA samples from control cultures for hybridization to six two-color microarrays. A dye-swap design was used to remove the Cy5 and Cy3 dye bias.
Project description:Yersinia pestis, the etiological agent of plague, is able to sense cell density by quorum sensing. The function of quorum sensing in Y. pestis is not clear. Here, the process of quorum sensing was investigated by comparing transcript profiles when three quorum-sensing signals are added in. The strain M-bM-^HM-^Fpgm (pigmentation-negative) mutant R88 was used as wild type. The three signals are AI-2, AHLs (N-(3-Oxooctanoyl)-L-homoserine lactone and N-Hexanoyl-DL-homoserine lactone).The control consisted of cells grown and treated under the same conditions without added signals. Six independent RNA samples from Y. pestis CO92 M-bM-^HM-^Fpgm cultures were paired with six independent RNA samples from 3 signals added cultures for hybridization to six two-color microarrays. A dye-swap design was used to remove the Cy5 and Cy3 dye bias.
Project description:The quorum-sensing system has been linked to diverse phenotypes and regulatory changes in pathogenic bacteria. In strain CO92, the AI-2 signal is produced in a luxS-dependent manner, reaching maximal levels of 2.5 μM in late logarithmic growth, and both wild type and pigmentation mutant strains made equivalent levels of AI-2. Yersinia pestis CO92 possesses a chromosomal lsr locus encoding factors involved in the binding and import of AI-2, and confirming this assignment, an lsr deletion increased extracellular pools of AI-2. To assess the functional role of 3 quorum sensing singlas in Y. pestis, microarray study was conducted comparing the 3 quorum sensing signals added in to control at 30°C to mimic the flea gut. Here, the ΔPgm (pigmentation-negative) R88 strain was used. The three signals are one AI-2 and two AHLs (N-(3-Oxooctanoyl)-L-homoserine lactone and N-Hexanoyl-DL-homoserine lactone).The control consisted of cells grown and treated under the same conditions without added signals.
Project description:Yersinia pestis, the etiological agent of plague, is able to sense cell density by quorum sensing. The function of quorum sensing in Y. pestis is not clear. Here, the process of AHL quorum sensing was investigated by comparing transcript profiles when two AHL quorum-sensing signals are added in. The strain Îpgm (pigmentation-negative) mutant was called wild type. The two AHLs signals are N-(3-Oxooctanoyl)-L-homoserine lactone and N-Hexanoyl-DL-homoserine lactone.The control consisted of cells grown and treated under the same conditions without added signals. Six independent RNA samples from Y. pestis CO92 Îpgm cultures were paired with six independent RNA samples from two AHLs added cultures for hybridization to six two-color microarrays. A dye-swap design was used to remove the Cy5 and Cy3 dye bias.
Project description:Yersinia pestis, the etiological agent of plague, is able to sense cell density by quorum sensing. The function of quorum sensing in Y. pestis is not clear. Here, the process of AHL quorum sensing was investigated by comparing transcript profiles when two AHL quorum-sensing signals are added in. The strain âpgm (pigmentation-negative) mutant R88 was called wild type. The two AHLs signals are N-(3-Oxooctanoyl)-L-homoserine lactone and N-Hexanoyl-DL-homoserine lactone.The control consisted of cells grown and treated under the same conditions without added signals. Six independent RNA samples from R88 cultures were paired with six independent RNA samples from two AHLs added cultures for hybridization to six two-color microarrays. A dye-swap design was used to remove the Cy5 and Cy3 dye bias.
Project description:We examined whether the budding yeast Saccharomyces cerevisiae can sense chemical signals from prokaryotes, specifically a variety of quorum sensing molecules from different bacteria species and from Candida albicans. We found that only N-acyl-3-oxo-dodecanoyl homoserine lactone (C12) from the opportunistic human pathogen Pseudomonas aeruginosa induces a stress response in yeast. Microarray experiments were performed in order to continue investigating the stress response. We treated S. cerevesiae (WT strain W303) with N-(3-oxo-dodecanoyl) homoserine lactone (C12), a quorum sensing molecule of Pseudomonas aeruginosa, which we found causes a stress response using a GFP reporter for HSP-12. Treatment conditions: 100 uM C12, 100 uM C12-lactam (control: synthetic analogue of C12 that is inactive in P. aeruginosa), DMSO (control: solvent), and 0.3 mM H2O2 (for comparison to oxidative stress).
Project description:Yersinia pestis, the etiological agent of plague, is able to sense cell density by quorum sensing. The function of quorum sensing in Y. pestis is not clear. Here, the process of quorum sensing was investigated by comparing transcript profiles when three quorum-sensing signals are added in. The strain ∆pgm (pigmentation-negative) mutant R88 was used as wild type. The three signals are AI-2, AHLs (N-(3-Oxooctanoyl)-L-homoserine lactone and N-Hexanoyl-DL-homoserine lactone).The control consisted of cells grown and treated under the same conditions without added signals.
Project description:The AI-2 quorum-sensing system has been linked to diverse phenotypes and regulatory changes in pathogenic bacteria. In strain CO92, the AI-2 signal is produced in a luxS-dependent manner, reaching maximal levels of 2.5 μM in late logarithmic growth, and both wild type and pigmentation mutant strains made equivalent levels of AI-2. Y. pestis CO92 possesses a chromosomal lsr locus encoding factors involved in the binding and import of AI-2, and confirming this assignment, an lsr deletion increased extracellular pools of AI-2. To assess the functional role of AI-2 sensing in Y. pestis, microarray study was conducted comparing the ∆Pgm strain R88 to a ∆Pgm ∆luxS mutant at 30°C to mimic the flea gut.
Project description:In order to gain coherent insights into plant responses we performed transcriptional analysis of Arabidopsis seedling after treatment with three different AHLs: N-hexanoyl-L-homoserine lactone (C6-HSL), N-3-oxo-decanoyl-L-homoserine lactone (oxo-C10-HSL), and N-3-oxo-tetradecanoyl-L-homoserine lactone (oxo-C14-HSL). Furthermore, we analyzed the transcriptome of oxo-C14-HSL pretreated plants after a secondary challenge with 100 nM flg22 for 2 and 24 hours after treatment. Gene expression in Arabidopsis thaliana seedlings were measured after a 3-days-pretreatment with 6 µM of three different N-acyl homoserine lactones in comparison to plants treated with the coresponding volume of acetone (solvent control). Plants pretreated with oxo-C14-HSL were in addition treated with 100nM of flg22 to induced defense response. Three independent experiments were performed (replicates).
Project description:The etiologic agent of bubonic plague, Yersinia pestis, senses cell density-dependent chemical signals to synchronize transcription between cells of the population in a process named quorum sensing. Though the closely related enteric pathogen Y. pseudotuberculosis uses quorum sensing system to regulate motility, the role of quorum sensing in Y. pestis has been unclear. In this study we performed transcriptional profiling experiments to identify Y. pestis quorum sensing regulated functions. Our analysis revealed that acyl-homoserine lactone based quorum sensing controls the expression of several metabolic functions. Maltose fermentation and the glyoxylate bypass are induced by acyl-homoserine lactone signaling. This effect was seen to be temperature conditional. Metabolism is unresponsive to quorum sensing regulation at mammalian body temperature, indicating a potential role for quorum sensing regulation of metabolism specifically during colonization of the flea vector. It is proposed that utilization of alternative carbon sources may enhance growth and/or survival during prolonged flea colonization, contributing to maintenance of plague in nature. Six independent RNA samples from Y. pestis CO92 R114 AHL deficient cultures were paired with six independent RNA samples from control Y. pestis CO92 R88 cultures for hybridization to six two-color microarrays. For three arrays, the control RNA sample was labeled with Alexa 555 dye and the experimental RNA sample was labeled with Alexa 647 dye; the dyes were reversed for the other three arrays to account for any dye bias.
Project description:The AI-2 quorum-sensing system has been linked to diverse phenotypes and regulatory changes in pathogenic bacteria. In strain CO92, the AI-2 signal is produced in a luxS-dependent manner, reaching maximal levels of 2.5 μM in late logarithmic growth, and both wild type and pigmentation mutant strains made equivalent levels of AI-2. Y. pestis CO92 possesses a chromosomal lsr locus encoding factors involved in the binding and import of AI-2, and confirming this assignment, an lsr deletion increased extracellular pools of AI-2. To assess the functional role of AI-2 sensing in Y. pestis, microarray study was conducted comparing the âPgm strain R88 to a âPgm âluxS mutant at 30°C to mimic the flea gut. Six independent RNA samples from Y. pestis CO92 ÎPgm cultures were paired with six independent RNA samples from ÎPgm ÎluxS cultures for hybridization to six two-color microarrays. A dye-swap design was used to remove the Cy5 and Cy3 dye bias.