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1.
Nat Commun ; 14(1): 7986, 2023 Dec 02.
Article in English | MEDLINE | ID: mdl-38042853

ABSTRACT

Quorum sensing is a mechanism of bacterial communication that controls virulence gene expression. Pseudomonas aeruginosa regulates virulence via two synthase/transcription factor receptor pairs: LasI/R and RhlI/R. LasR is considered the master transcriptional regulator of quorum sensing, as it upregulates rhlI/R. However, clinical isolates often have inactivating mutations in lasR, while maintaining Rhl-dependent signaling. We sought to understand how quorum sensing progresses in isolates with lasR mutations, specifically via activation of RhlR. We find that clinical isolates with lasR inactivating mutations often harbor concurrent mutations in rhlI. Using ultra-high-performance liquid chromatography coupled with high-resolution mass spectrometry, we discover that strains lacking lasR overproduce the RhlI-synthesized autoinducer and that RhlI variants re-calibrate autoinducer concentrations to wild-type levels, restoring virulent phenotypes. These findings provide a mechanism for the plasticity of quorum sensing progression in an acute infection niche.


Subject(s)
Pseudomonas aeruginosa , Trans-Activators , Trans-Activators/metabolism , Pseudomonas aeruginosa/metabolism , Quorum Sensing/genetics , Mutation , Transcription Factors/metabolism , Nitric Oxide Synthase/metabolism , Bacterial Proteins/metabolism , Gene Expression Regulation, Bacterial
2.
Structure ; 30(12): 1626-1636.e4, 2022 12 01.
Article in English | MEDLINE | ID: mdl-36379213

ABSTRACT

Pseudomonas aeruginosa is an opportunistic pathogen that is responsible for thousands of deaths every year in the United States. P. aeruginosa virulence factor production is mediated by quorum sensing, a mechanism of bacterial cell-cell communication that relies on the production and detection of signal molecules called autoinducers. In P. aeruginosa, the transcription factor receptor RhlR is activated by a RhlI-synthesized autoinducer. We recently showed that RhlR-dependent transcription is enhanced by a physical interaction with the enzyme PqsE via increased affinity of RhlR for promoter DNA. However, the molecular basis for complex formation and how complex formation enhanced RhlR transcriptional activity remained unclear. Here, we report the structure of ligand-bound RhlR in complex with PqsE. Additionally, we determined the structure of the complex bound with DNA, revealing the mechanism by which RhlR-mediated transcription is enhanced by PqsE, thereby establishing the molecular basis for RhlR-dependent virulence factor production in P. aeruginosa.


Subject(s)
Pseudomonas aeruginosa , Quorum Sensing , Quorum Sensing/genetics , Pseudomonas aeruginosa/genetics , Pseudomonas aeruginosa/metabolism , Bacterial Proteins/genetics , Bacterial Proteins/metabolism , Gene Expression Regulation, Bacterial , Virulence Factors/genetics , Virulence Factors/metabolism
3.
Microorganisms ; 10(6)2022 Jun 18.
Article in English | MEDLINE | ID: mdl-35744765

ABSTRACT

Bacteria use a cell-cell communication process called quorum sensing (QS) to orchestrate collective behaviors. QS relies on the group-wide detection of extracellular signal molecules called autoinducers (AI). Quorum sensing is required for virulence and biofilm formation in the human pathogen Pseudomonas aeruginosa. In P. aeruginosa, LasR and RhlR are homologous LuxR-type soluble transcription factor receptors that bind their cognate AIs and activate the expression of genes encoding functions required for virulence and biofilm formation. While some bacterial signal transduction pathways follow a linear circuit, as phosphoryl groups are passed from one carrier protein to another ultimately resulting in up- or down-regulation of target genes, the QS system in P. aeruginosa is a dense network of receptors and regulators with interconnecting regulatory systems and outputs. Once activated, it is not understood how LasR and RhlR establish their signaling hierarchy, nor is it clear how these pathway connections are regulated, resulting in chronic infection. Here, we reviewed the mechanisms of QS progression as it relates to bacterial pathogenesis and antimicrobial resistance and tolerance.

4.
Microbiol Spectr ; 10(1): e0210821, 2022 02 23.
Article in English | MEDLINE | ID: mdl-35019777

ABSTRACT

Pseudomonas aeruginosa is an opportunistic pathogen that causes disease in immunocompromised individuals and individuals with underlying pulmonary disorders. P. aeruginosa virulence is controlled by quorum sensing (QS), a bacterial cell-cell communication mechanism that underpins transitions between individual and group behaviors. In P. aeruginosa, the PqsE enzyme and the QS receptor RhlR directly interact to control the expression of genes involved in virulence. Here, we show that three surface-exposed arginine residues on PqsE comprise the site required for interaction with RhlR. We show that a noninteracting PqsE variant [PqsE(NI)] possesses catalytic activity, but is incapable of promoting virulence phenotypes, indicating that interaction with RhlR, and not catalysis, drives these PqsE-dependent behaviors. Biochemical characterization of the PqsE-RhlR interaction coupled with RNA-seq analyses demonstrates that the PqsE-RhlR complex increases the affinity of RhlR for DNA, enabling enhanced expression of genes encoding key virulence factors. These findings provide the mechanism for PqsE-dependent regulation of RhlR and identify a unique regulatory feature of P. aeruginosa QS and its connection to virulence. IMPORTANCE Bacteria use a cell-cell communication process called quorum sensing (QS) to orchestrate collective behaviors. QS relies on the group-wide detection of molecules called autoinducers (AI). QS is required for virulence in the human pathogen Pseudomonas aeruginosa, which can cause fatal infections in patients with underlying pulmonary disorders. In this study, we determine the molecular basis for the physical interaction between two virulence-driving QS components, PqsE and RhlR. We find that the ability of PqsE to bind RhlR correlates with virulence factor production. Since current antimicrobial therapies exacerbate the growing antibiotic resistance problem because they target bacterial growth, we suggest that the PqsE-RhlR interface discovered here represents a new candidate for targeting with small molecule inhibition. Therapeutics that disrupt the PqsE-RhlR interaction should suppress virulence. Targeting bacterial behaviors such as QS, rather than bacterial growth, represents an attractive alternative for exploration because such therapies could potentially minimize the development of resistance.


Subject(s)
Bacterial Proteins/metabolism , DNA-Binding Proteins/metabolism , Pseudomonas aeruginosa/metabolism , Virulence Factors/metabolism , Anti-Bacterial Agents/pharmacology , Bacterial Proteins/chemistry , Bacterial Proteins/genetics , Cell Communication/drug effects , DNA-Binding Proteins/chemistry , DNA-Binding Proteins/genetics , Gene Expression Regulation, Bacterial/drug effects , Humans , Pseudomonas aeruginosa/genetics , Quorum Sensing/physiology , Virulence , Virulence Factors/genetics
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