Project description:Hafnia alvei H4 is a bacteria subject to regulation by N-acyl-l-homoserine lactone (AHL)-mediated quorum sensing system and is closely related to the corruption of instant sea cucumber. Studying the effect of Hafnia alvei H4 quorum sensing regulatory genes on AHLs is necessary for the quality and preservation of instant sea cucumber. In this study, the draft genome of H. alvei H4, which comprises a single chromosome of 4,687,151 bp, was sequenced and analyzed and the types of AHLs were analyzed employing thin-layer chromatography (TLC) and LC-MS/MS. Then the wild-type strain of H. alvei H4 and the luxI/R double mutant (ΔluxIR) were compared by transcriptome sequencing (RNA-seq). The results indicate that the incomplete genome sequence revealed the presence of one quorum-sensing (QS) gene set, designated as lasI/expR. Three major AHLs, N-hexanoyl-L-homoserine lactone (C6-HSL), N-butyryl-L-homoserine lactone (C4-HSL), and N-(3-oxo-octanoyl)-L-homoserine lactone (3-oxo-C8-HSL) were found, with C6-HSL being the most abundant. C6-HSL was not detected in the culture of the luxI mutant (ΔluxI) and higher levels of C4-HSL was found in the culture of the luxR mutant (ΔluxR), which suggested that the luxR gene may have a positive effect on C4-HSL production. It was also found that AHL and QS genes are closely related in the absence of luxIR double deletion. The results of this study can further elucidate at the genetic level that luxI and luxR genes are involved in the regulation of AHL.
Project description:Analysis of the effect of a bacterial quorum sensing molecule on in-vitro culture of human endothelial cells at gene expression levels. Objective: Chronic infection has long been postulated as a stimulus for atherogenesis. Pseudomonas aeruginosa infection has been associated with increased atherosclerosis in rats, and the bacteria produce a quorum sensing molecule 3-oxo-dodecynoyl-homoserine lactone (3OC12-HSL) that is critical for colonization and virulence. Paraoxonase 2 (PON2) hydrolyzes 3OC12-HSL and also protects against the effects of oxidized phospholipids thought to contribute to atherosclerosis. We now report the response of human aortic endothelial cells (HAEC) to 3OC12-HSL and oxidized 1-palmitoyl-2-arachidonoyl-sn-glycero-3-phosphocholine (Ox-PAPC) in relation to PON2 expression. Methods and Results: Using expression profiling and network modeling, we identified the unfolded protein response (UPR), cell cycle genes, and the MAPK signaling pathway to be heavily involved in the HAEC response to 3OC12-HSL. The network also showed striking similarities to a network created based on HAEC response to Ox-PAPC, a major component of minimally-modified LDL. HAEC in which PON2 was silenced by siRNA showed increased pro-inflammatory and UPR responses when treated with 3OC12-HSL or Ox-PAPC. Conclusion: 3OC12-HSL and Ox-PAPC influence similar inflammatory pathways. Quorum sensing molecules such as 3OC12-HSL contribute to the pro-atherogenic effects of chronic infection and the anti-atherogenic effects of PON2 include destruction of quorum sensing molecules. 4 HAEC lines from different donors were treated with 3-O-C12 HSL or control (medium).
Project description:Analysis of the effect of a bacterial quorum sensing molecule on in-vitro culture of human endothelial cells at gene expression levels. Objective: Chronic infection has long been postulated as a stimulus for atherogenesis. Pseudomonas aeruginosa infection has been associated with increased atherosclerosis in rats, and the bacteria produce a quorum sensing molecule 3-oxo-dodecynoyl-homoserine lactone (3OC12-HSL) that is critical for colonization and virulence. Paraoxonase 2 (PON2) hydrolyzes 3OC12-HSL and also protects against the effects of oxidized phospholipids thought to contribute to atherosclerosis. We now report the response of human aortic endothelial cells (HAEC) to 3OC12-HSL and oxidized 1-palmitoyl-2-arachidonoyl-sn-glycero-3-phosphocholine (Ox-PAPC) in relation to PON2 expression. Methods and Results: Using expression profiling and network modeling, we identified the unfolded protein response (UPR), cell cycle genes, and the MAPK signaling pathway to be heavily involved in the HAEC response to 3OC12-HSL. The network also showed striking similarities to a network created based on HAEC response to Ox-PAPC, a major component of minimally-modified LDL. HAEC in which PON2 was silenced by siRNA showed increased pro-inflammatory and UPR responses when treated with 3OC12-HSL or Ox-PAPC. Conclusion: 3OC12-HSL and Ox-PAPC influence similar inflammatory pathways. Quorum sensing molecules such as 3OC12-HSL contribute to the pro-atherogenic effects of chronic infection and the anti-atherogenic effects of PON2 include destruction of quorum sensing molecules.
Project description:Quorum sensing is a term used to describe cell-to-cell communication that allows cell density-dependent gene expression. Many Gram-negative bacteria use acyl-homoserine lactone (acyl-HSL) synthases to generate fatty acyl-HSL quorum sensing signals, which function with signal receptors to control expression of specific genes. The fatty acyl group is derived from fatty acid biosynthesis and provides signal specificity, but the variety of signals is limited. We have discovered that the photosynthetic bacterium Rhodopseudomonas palustris uses an acyl-HSL synthase to produce p-coumaroyl-HSL by using environmental p-coumaric acid rather than fatty acids from cellular pools. The bacterium has a signal receptor with homology to fatty acyl-HSL receptors that responds to p-coumaroyl-HSL to regulate global gene expression. We also found that p-coumaroyl-HSL is made by other bacteria including Bradyrhizobium BTAi1 and Silicibacter pomeroyi DSS-3. This discovery extends the range of possibilities for acyl-HSL quorum sensing and raises fundamental questions about quorum sensing within the context of environmental signaling. Keywords: Comparison of transcriptome profiles Transcriptome profiles between Rhodopseudomonas palustris cells grown in the in the presence or absence of pC-HSL were compared.
Project description:Quorum sensing is a term used to describe cell-to-cell communication that allows cell density-dependent gene expression. Many Gram-negative bacteria use acyl-homoserine lactone (acyl-HSL) synthases to generate fatty acyl-HSL quorum sensing signals, which function with signal receptors to control expression of specific genes. The fatty acyl group is derived from fatty acid biosynthesis and provides signal specificity, but the variety of signals is limited. We have discovered that the photosynthetic bacterium Rhodopseudomonas palustris uses an acyl-HSL synthase to produce p-coumaroyl-HSL by using environmental p-coumaric acid rather than fatty acids from cellular pools. The bacterium has a signal receptor with homology to fatty acyl-HSL receptors that responds to p-coumaroyl-HSL to regulate global gene expression. We also found that p-coumaroyl-HSL is made by other bacteria including Bradyrhizobium BTAi1 and Silicibacter pomeroyi DSS-3. This discovery extends the range of possibilities for acyl-HSL quorum sensing and raises fundamental questions about quorum sensing within the context of environmental signaling. Keywords: Comparison of transcriptome profiles
Project description:Acyl-homoserine lactone (acyl-HSL) quorum sensing was first discovered in Vibrio fischeri where it serves as a key control element of the seven-gene luminescence (lux) operon. Since this initial discovery, other bacteria have been shown to control hundreds of genes by acyl-HSL quorum sensing. Until recently, it has been difficult to examine the global nature of quorum sensing in V. fischeri. However, the complete genome sequence of V. fischeri is now available and this has enabled us to use transcriptomics to identify quorum-sensing regulated genes and to study the quorum-controlled regulon of this bacterium. In this study, we used DNA microarray technology to identify over two-dozen V. fischeri genes regulated by the quorum sensing signal N-3-oxohexanoyl-L-homoserine lactone (3OC6-HSL). Keywords: Comparison of transcriptome profiles
Project description:The human opportunistic pathogen Pseudomonas aeruginosa orchestrates the expression of many genes in a cell density-dependent manner by using a molecular communication system referred to as quorum sensing (QS). This bacterium uses an intricate network of regulators and QS molecules known as autoinducers. Among these autoinducers are two acyl-homoserine lactones (AHL) involved in QS circuits which modulate virulence factors production, biofilm formation, and antimicrobial sensitivity. Disrupting QS, a strategy referred to as quorum quenching (QQ), is a promising approach to modulate virulence while not directly challenging bacterial survival. For P. aeruginosa, QQ can be achieved using exogenous AHL-degrading lactonases. However, the importance of enzyme specificity on quenching efficacy has never been investigated. Here, we used two lactonases both targeting the signal molecules N-(3-oxododecanoyl)-L-Homoserine lactone (3-oxo-C12 HSL) and butyryl-homoserine lactone (C4 HSL) albeit with different efficacy on C4 HSL. Interestingly, both lactonases similarly decreased the concentrations of AHL and comparably impacted the expression of AHL-based circuits. Conversely, strong variations were observed in Pseudomonas Quinolone Signal (PQS) regulation. Both lactonases were then found to decrease virulence factors production and biofilm formation in vitro, albeit with different efficiencies. Unexpectedly, only the lactonase with substrate preference for 3-oxo-C12 HSL was able to inhibit P. aeruginosa pathogenicity in vivo in an amoeba infection model. Similarly, large variations between lactonases were observed in proteins involved in antibiotic resistance, biofilm formation, virulence and diverse cellular mechanisms. This global analysis provides the first evidence that QQ enzyme specificity is crucial to modulate QS-associated behavior in P. aeruginosa PA14.
2020-05-27 | PXD017421 | Pride
Project description:Impact of C4-HSL-mediated quorum sensing on acidogenic fermentation under saline conditions
Project description:Quorum-sensing (QS) is a cell-cell communication system that controls gene expression in many bacterial species, mediated by diffusible signal molecules. While the intracellular regulatory mechanisms of QS are often well-understood, the functional roles of QS remain controversial. In particular, the use of multiple signals by many bacterial species poses a serious challenge to current functional theories. Here we address this challenge by showing that bacteria can use multiple QS signals to infer both their social (density) and physical (mass-transfer) environment. Analytical and evolutionary simulation models show that the detection of and response to complex social/physical contrasts requires multiple signals with distinct half-lives and combinatorial (non-additive) responses to signal concentrations. We test these predictions using the opportunistic pathogen Pseudomonas aeruginosa, and demonstrate significant differences in signal decay between its two primary signal molecules as well as diverse combinatorial responses to dual signal inputs. QS is associated with the control of secreted factors, and we show that secretome genes are preferentially controlled by synergistic M-bM-^@M-^XAND-gateM-bM-^@M-^Y responses to multiple signal inputs, ensuring the effective expression of secreted factors in high density and low mass-transfer environments. Our results support a novel functional hypothesis for the use of multiple signals and, more generally, show that bacteria are capable of combinatorial communication. The two primary signal molecules of P. aeruginosa are the homoserine lactones N-(3-oxododecanoyl)-L-homoserine lactone (3-oxo-C12-HSL) and N-butyryl-homoserine lactone (C4-HSL). Effects of the different signal molecules was assessed using a double QS synthase mutant of Pseudomonas aeruginosa PAO1 lasI/rhlI grown at 37M-BM-0C in 25 ml LB broth and 250 ml flasks with shaking at 200 r.p.m. in four treatments, each with a replicate: (a) no addition; (b) 3-oxo- C12-HSL; (c) C4-HSL; and (d) both 3-oxo-C12-HSL and C4-HSL.
Project description:Comparison of a quorum sensing null mutant supplemented with C18-HSL, C18en-HSL, C18dien-HSL with D. shibae wild-type in order to identify traits induced by distinct autoinducers in this organism.