Chemical inhibitors of the conserved bacterial transcriptional regulator DksA1 suppressed quorum sensing-mediated virulence of Pseudomonas aeruginosa

Pseudomonas aeruginosa uses the two major quorum-sensing (QS) regulatory systems las and rhl to modulate the expression of many of its virulence factors. The las system is considered to stand at the top of the QS hierarchy. However, some virulence factors such as pyocyanin have been reported to still be produced in lasR mutants under certain conditions. Interestingly, such mutants arise spontaneously under various conditions, including in the airways of cystic fibrosis patients. Using transcriptional lacZ reporters, LC/MS quantification and phenotypic assays, we have investigated the regulation of QS-controlled factors by the las system. Our results show that activity of the rhl system is only delayed in a lasR mutant, thus allowing the expression of multiple virulence determinants such as pyocyanin, rhamnolipids and C4-homoserine lactone (HSL) during the late stationary phase. Moreover, at this stage, RhlR is able to overcome the absence of the las system by activating specific LasR-controlled functions, including production of 3-oxo-C12-HSL and Pseudomonas quinolone signal (PQS). P. aeruginosa is thus able to circumvent the deficiency of one of its QS systems by allowing the other to take over. This work demonstrates that the QS hierarchy is more complex than the model simply presenting the las system above the rhl system.

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Environmental Regulation of Pseudomonas aeruginosa PAO1 Las and Rhl Quorum-Sensing Systems

Journal of Bacteriology ◽

10.1128/jb.00043-07 ◽

2007 ◽

Vol 189 (13) ◽

pp. 4827-4836 ◽

Cited By ~ 146

Author(s):

Kangmin Duan ◽

Michael G. Surette

Keyword(s):

Pseudomonas Aeruginosa ◽

Quorum Sensing ◽

Environmental Conditions ◽

Transcriptional Regulator ◽

Transcriptional Regulators ◽

Growth Conditions ◽

Fine Tuning ◽

Relative Timing ◽

Sensing Systems ◽

Two Systems

ABSTRACT The lasI-lasR and the rhlI-rhlR quorum-sensing systems in Pseudomonas aeruginosa regulate the expression of numerous cellular and secreted virulence factor genes and play important roles in the development of biofilms. The las and rhl systems themselves are known to be directly or indirectly regulated by a number of transcriptional regulators, and consequently, their expression is sensitive to environmental conditions. In this report, the activities of these two quorum-sensing systems have been examined systematically under 46 growth conditions, and the regulation by environmental conditions has been investigated. The relative timing and strength of expression of these two systems varied significantly under different conditions, which contrasts with the notion of a preset hierarchy with these two systems in P. aeruginosa. Depending on the growth conditions, the correlation between each synthase and its cognate transcriptional regulator also varied, suggesting that the transcription of these genes independently allows for further fine tuning of each system. Finally, we observe that the activities of both the lasI-lasR and the rhlI-rhlR quorum-sensing systems were dramatically enhanced in the presence of extracts of sputum samples from cystic fibrosis patients.

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The Pseudomonas aeruginosa rmlBDAC operon, encoding dTDP-l-rhamnose biosynthetic enzymes, is regulated by the quorum-sensing transcriptional regulator RhlR and the alternative sigma factor σS

Microbiology ◽

10.1099/mic.0.054726-0 ◽

2012 ◽

Vol 158 (4) ◽

pp. 908-916 ◽

Cited By ~ 29

Author(s):

Marisela Aguirre-Ramírez ◽

Gerardo Medina ◽

Abigail González-Valdez ◽

Victoria Grosso-Becerra ◽

Gloria Soberón-Chávez

Keyword(s):

Pseudomonas Aeruginosa ◽

Quorum Sensing ◽

Sigma Factor ◽

Transcriptional Regulator ◽

Alternative Sigma Factor ◽

Biosynthetic Enzymes

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Interaction of quercetin with transcriptional regulator LasR of Pseudomonas aeruginosa: Mechanistic insights of the inhibition of virulence through quorum sensing

10.1101/239996 ◽

2017 ◽

Author(s):

Hovakim Grabski ◽

Lernik Hunanyan ◽

Susanna Tiratsuyan ◽

Hrachik Vardapetyan

Keyword(s):

Molecular Dynamics ◽

Pseudomonas Aeruginosa ◽

Quorum Sensing ◽

Biofilm Formation ◽

Principal Component ◽

Transcriptional Regulator ◽

Binding Mode ◽

World Health ◽

Hydroxyl Group ◽

Binding Modes

ABSTRACTBackgroundPseudomonas aeruginosais one of the most dangerous superbugs in the list of bacteria for which new antibiotics are urgently needed, which was published by World Health Organization.P. aeruginosais an antibiotic-resistant opportunistic human pathogen. It affects patients with AIDS, cystic fibrosis, cancer, burn victims and people with prosthetics and implants.P. aeruginosaalso forms biofilms. Biofilms increase resistance to antibiotics and host immune responses. Because of biofilms, current therapies are not effective. It is important to find new antibacterial treatment strategies againstP. aeruginosa. Biofilm formation is regulated through a system called quorum sensing. Thus disrupting this system is considered a promising strategy to combat bacterial pathogenicity. It is known that quercetin inhibitsPseudomonas aeruginosabiofilm formation, but the mechanism of action is unknown. In the present study, we tried to analyse the mode of interactions of LasR with quercetin.ResultsWe used a combination of molecular docking, molecular dynamics (MD) simulations and machine learning techniques for the study of the interaction of the LasR protein ofP. aeruginosawith quercetin. We assessed the conformational changes of the interaction and analysed the molecular details of the binding of quercetin with LasR. We show that quercetin has two binding modes. One binding mode is the interaction with ligand binding domain, this interaction is not competitive and it has also been shown experimentally. The second binding mode is the interaction with the bridge, it involves conservative amino acid interactions from LBD, SLR, and DBD and it is also not competitive. Experimental studies show hydroxyl group of ring A is necessary for inhibitory activity, in our model the hydroxyl group interacts with Leu177 during the second binding mode. This could explain the molecular mechanism of how quercetin inhibits LasR protein.ConclusionsThis study may offer insights on how quercetin inhibits quorum sensing circuitry by interacting with transcriptional regulator LasR. The capability of having two binding modes may explain why quercetin is effective at inhibiting biofilm formation and virulence gene expression.List of abbreviationsPDBProtein data bankMDMolecular DynamicsPCAPrincipal Component AnalysisPCPrincipal ComponentSLRShort Linker RegionBLASTBasic local alignment search toolDBIDavid-Bouldin IndexpsFpseudo-F statistic

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Functions Required for Extracellular Quinolone Signaling by Pseudomonas aeruginosa

Journal of Bacteriology ◽

10.1128/jb.184.23.6472-6480.2002 ◽

2002 ◽

Vol 184 (23) ◽

pp. 6472-6480 ◽

Cited By ~ 372

Author(s):

Larry A. Gallagher ◽

Susan L. McKnight ◽

Marina S. Kuznetsova ◽

Everett C. Pesci ◽

Colin Manoil

Keyword(s):

Pseudomonas Aeruginosa ◽

Quorum Sensing ◽

Secondary Metabolites ◽

Regulatory Network ◽

Biosynthetic Pathway ◽

Extracellular Enzymes ◽

Transcriptional Regulator ◽

Transposon Mutagenesis ◽

Sensing System ◽

Quorum Sensing System

ABSTRACT A set of 30 mutants exhibiting reduced production of the phenazine poison pyocyanin were isolated following transposon mutagenesis of Pseudomonas aeruginosa PAO1. The mutants could be subdivided into those with defects in the primary phenazine biosynthetic pathway and those with more pleiotropic defects. The largest set of pleiotropic mutations blocked the production of the extracellular Pseudomonas quinolone signal (PQS), a molecule required for the synthesis of secondary metabolites and extracellular enzymes. Most of these pqs mutations affected genes which appear to encode PQS biosynthetic functions, although a transcriptional regulator and an apparent response effector were also represented. Two of the genes required for PQS synthesis (phnA and phnB) had previously been assumed to encode phenazine biosynthetic functions. The transcription of one of the genes required for PQS synthesis (PA2587/pqsH) was regulated by the LasI/R quorum-sensing system, thereby linking quorum sensing and PQS regulation. Others of the pleiotropic phenazine-minus mutations appear to inactivate novel components of the quorum-sensing regulatory network, including one regulator (np20) previously shown to be required for virulence in neutropenic mice.

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PqsE Is Essential for RhlR-Dependent Quorum Sensing Regulation in Pseudomonas aeruginosa

mSystems ◽

10.1128/msystems.00194-20 ◽

2020 ◽

Vol 5 (3) ◽

Author(s):

Marie-Christine Groleau ◽

Thays de Oliveira Pereira ◽

Valérie Dekimpe ◽

Eric Déziel

Keyword(s):

Pseudomonas Aeruginosa ◽

Quorum Sensing ◽

Cell Communication ◽

Transcriptional Regulator ◽

Homoserine Lactone ◽

World Health ◽

Chronic Infections ◽

Content Type ◽

Main Regulator ◽

Health Organization

ABSTRACT The bacterium Pseudomonas aeruginosa has emerged as a central threat in health care settings and can cause a large variety of infections. It expresses an arsenal of virulence factors and a diversity of survival functions, many of which are finely and tightly regulated by an intricate circuitry of three quorum sensing (QS) systems. The las system is considered at the top of the QS hierarchy and activates the rhl and pqs systems. It is composed of the LasR transcriptional regulator and the LasI autoinducer synthase, which produces 3-oxo-C12-homoserine lactone (3-oxo-C12-HSL), the ligand of LasR. RhlR is the transcriptional regulator for the rhl system and is associated with RhlI, which produces its cognate autoinducer C4-HSL. The third QS system is composed of the pqsABCDE operon and the MvfR (PqsR) regulator. PqsABCD synthetize 4-hydroxy-2-alkylquinolines (HAQs), which include ligands activating MvfR. PqsE is not required for HAQ production and instead is associated with the expression of genes controlled by the rhl system. While RhlR is often considered the main regulator of rhlI, we confirmed that LasR is in fact the principal regulator of C4-HSL production and that RhlR regulates rhlI and production of C4-HSL essentially only in the absence of LasR by using liquid chromatography-mass spectrometry quantifications and gene expression reporters. Investigating the expression of RhlR targets also clarified that activation of RhlR-dependent QS relies on PqsE, especially when LasR is not functional. This work positions RhlR as the key QS regulator and points to PqsE as an essential effector for full activation of this regulation. IMPORTANCE Pseudomonas aeruginosa is a versatile bacterium found in various environments. It can cause severe infections in immunocompromised patients and naturally resists many antibiotics. The World Health Organization listed it among the top priority pathogens for research and development of new antimicrobial compounds. Quorum sensing (QS) is a cell-cell communication mechanism, which is important for P. aeruginosa adaptation and pathogenesis. Here, we validate the central role of the PqsE protein in QS particularly by its impact on the regulator RhlR. This study challenges the traditional dogmas of QS regulation in P. aeruginosa and ties loose ends in our understanding of the traditional QS circuit by confirming RhlR to be the main QS regulator in P. aeruginosa. PqsE could represent an ideal target for the development of new control methods against the virulence of P. aeruginosa. This is especially important when considering that LasR-defective mutants frequently arise, e.g., in chronic infections.

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Quorum-Sensing Regulation of a Copper Toxicity System in Pseudomonas aeruginosa

Journal of Bacteriology ◽

10.1128/jb.01528-09 ◽

2010 ◽

Vol 192 (10) ◽

pp. 2557-2568 ◽

Cited By ~ 29

Author(s):

Joshua T. Thaden ◽

Stephen Lory ◽

Timothy S. Gardner

Keyword(s):

Pseudomonas Aeruginosa ◽

Quorum Sensing ◽

Molecular Genetic ◽

Copper Toxicity ◽

Transcriptional Regulator ◽

Global Gene Expression ◽

Copper Resistance ◽

Binding Motif ◽

Sensing System ◽

Quorum Sensing System

ABSTRACT The LasR/LasI quorum-sensing system in Pseudomonas aeruginosa influences global gene expression and mediates pathogenesis. In this study, we show that the quorum-sensing system activates, via the transcriptional regulator PA4778, a copper resistance system composed of 11 genes. The quorum-sensing global regulator LasR was recently shown to directly activate transcription of PA4778, a cueR homolog and a MerR-type transcriptional regulator. Using molecular genetic methods and bioinformatics, we verify the interaction of LasR with the PA4778 promoter and further demonstrate the LasR binding site. We also identify a putative PA4778 binding motif and show that the protein directly binds to and activates five promoters controlling the expression of 11 genes—PA3519 to -15, PA3520, mexPQ-opmE, PA3574.1, and cueA, a virulence factor in a murine model. Using gene disruptions, we show that PA4778, along with 7 of 11 gene targets of PA4778, increases the sensitivity of P. aeruginosa to elevated copper concentrations. This work identifies a cellular function for PA4778 and four other previously unannotated genes (PA3515, PA3516, PA3517, and PA3518) and suggests a potential role for copper in the quorum response. We propose to name PA4778 cueR.

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Faculty Opinions recommendation of Revisiting the quorum-sensing hierarchy in Pseudomonas aeruginosa: the transcriptional regulator RhlR regulates LasR-specific factors.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.1162290.622778 ◽

2009 ◽

Author(s):

Fergal O'Gara

Keyword(s):

Pseudomonas Aeruginosa ◽

Quorum Sensing ◽

Transcriptional Regulator ◽

Specific Factors

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Pseudomonas aeruginosa isolates defective in function of the LasR quorum sensing regulator are frequent in diverse environmental niches

10.1101/2021.03.25.437011 ◽

2021 ◽

Author(s):

Marie-Christine Groleau ◽

Hélène Taillefer ◽

Antony T. Vincent ◽

Philippe Constant ◽

Eric Déziel

Keyword(s):

Pseudomonas Aeruginosa ◽

Quorum Sensing ◽

Contaminated Soils ◽

Transcriptional Regulator ◽

Opportunistic Pathogen ◽

Sensing Systems ◽

Niche Adaptation ◽

High Prevalence ◽

Full Activation

ABSTRACTThe saprophyte Pseudomonas aeruginosa is a versatile opportunistic pathogen causing infections in immunocompromised individuals. To facilitate its adaptation to a large variety of niches, this bacterium exploits population density-dependant gene regulation systems called quorum sensing. In P. aeruginosa, three distinct but interrelated quorum sensing systems (las, rhl and pqs) regulate the production of many survival and virulence functions. In prototypical strains, the las system, through its transcriptional regulator LasR, is important for the full activation of the rhl and pqs systems. Still, LasR-deficient isolates have been reported, mostly sampled from the lungs of people with cystic fibrosis, where they are considered selected by the chronic infection environment. In this study, we show that a defect in LasR activity appears to be an actually widespread mechanism of adaptation in this bacterium. Indeed, we found abundant LasR-defective isolates sampled from hydrocarbon-contaminated soils, hospital sink drains, and meat/fish market environments, using an approach based on phenotypic profiling, supported by gene sequencing. Interestingly, several LasR-defective isolates maintain an active rhl system or are deficient in pqs system signaling. The high prevalence of a LasR-defective phenotype among environmental P. aeruginosa isolates questions the role of quorum sensing in niche adaptation.

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