Triggering the ExoS regulon of Pseudomonas aeruginosa: A GFP-reporter analysis of exoenzyme (Exo) S, ExoT and ExoU synthesis

Hornef, Mathias W.; Roggenkamp, Andreas; Geiger, Anna Maria; Hogardt, Michael; Jacobi, Christoph A.; Heesemann, Jürgen

doi:10.1006/mpat.2000.0398

Cited by 48 publications

(54 citation statements)

References 43 publications

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“…Small changes in ExsA, ExsD, ExsC, or ExsE levels lead to large changes in T3SS expression, suggesting that this regulatory cascade may mediate bistable expression of T3SS. Single-cell studies confirm that not all P. aeruginosa bacteria induce expression of T3SS proteins in response to host cell contact or low-calcium cues, resulting in phenotypic heterogeneity within a clonal population (8)(9)(10).…”

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confidence: 93%

Cheating by type 3 secretion system-negative Pseudomonas aeruginosa during pulmonary infection

Czechowska¹,

McKeithen-Mead

Moussawi³

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

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Significance Bacterial pathogens frequently use type 3 secretion systems (T3SS) to counteract host immune responses to infection. T3SS expression is associated with increased virulence of Pseudomonas aeruginosa in humans and in animal models, but T3SS-negative bacteria often are isolated from acutely and chronically infected patients. We tested whether T3SS-negative bacteria could “cheat” during mixed infections with T3SS-positive bacteria in a murine model of acute pneumonia. Bacterial cheating occurred in the inflamed lung but only when T3SS-positive bacteria secreted the phospholipase A 2 effector, Exotoxin U. Phenotypically T3SS-expressing and –non-expressing bacteria co-exist within P. aeruginosa populations, suggesting that bacterial cheating might allow T3SS-negative organisms to establish themselves within a host.

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confidence: 93%

Cheating by type 3 secretion system-negative Pseudomonas aeruginosa during pulmonary infection

Czechowska¹,

McKeithen-Mead

Moussawi³

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

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“…Each of these systems possesses a homoserine lactone synthase (LasI or RhlI, respectively), a regulator protein (LasR or RhlR, respectively) that modulates gene transcription, and their cognate activators, N-(3-oxododecanoyl)-L-homoserine lactone (C12-HSL) and N-butanoyl-L-homoserine lactone (C4-HSL). Moreover, the P. aeruginosa genome encodes a type III secretion system (TTSS) similar to that found on the virulence plasmid (Yahr & Frank, 1994;Yahr et al, 1995Yahr et al, , 1996Hornef et al, 2000).Despite this capacity to produce an arsenal of multiple exoproduct virulence determinants P. aeruginosa is able to grow in biofilms which also exist in the lungs of CF patients (Singh et al, 2000;Costerton, 2001 De Kievit et al, 2001). As the exoenzyme S regulon is triggered by eukaryotic cell contact, we assume that pseudomonads embedded in bacterial biofilms should not exhibit a remarkable activity of type III effectors, which primarily are used to provoke eukaryotic cell intoxication.…”

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confidence: 99%

“…Prior to ExoS9-GFP measurement, bacterial cells were washed twice with PBS and finally resuspended in 1 ml PBS. P. aeruginosa PAO1 harbouring the control plasmid pKT-gfp (pKT248 carrying the gfp gene without the exoS promoter) was used as negative control (Hornef et al, 2000). A Coulter Epics flow cytometer (Beckman Coulter) equipped with an argon 488 nm laser was used to measure the intensity of fluorescence of ExoS9-GFP-producing bacteria.…”

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confidence: 99%

Expression of Pseudomonas aeruginosa exoS is controlled by quorum sensing and RpoS

et al. 2004

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In Pseudomonas aeruginosa, virulence determinants and biofilm formation are coordinated via a hierarchical quorum sensing cascade, which involves the transcriptional regulators LasR and RhlR and their cognate homoserine lactone activators C12-HSL [N-(3-oxododecanoyl)-L-homoserine lactone] and C4-HSL (N-butanoyl-L-homoserine lactone), which are produced by LasI and RhlI, respectively. The exoenzyme S regulon of P. aeruginosa, comprises genes for a type III secretion system and for four anti-host effector proteins (ExoS, T, U and Y), which are translocated into host cells. It is a reasonable assumption that this ExoS regulon should be downregulated in the biofilm growth state and thus should also be under the regulatory control of the Las/Rhl system. Therefore, an exoS9-gfp reporter construct was used, and the influence of the Las and Rhl quorum sensing systems and the effect of the stationary-phase sigma factor RpoS on regulation of the exoS gene was examined. Evidence is provided for downregulation of exoS during biofilm formation of P. aeruginosa PAO1. The rhlI mutant PDO100 and rhlR mutant PDO111, but not the lasI mutant PDO-JP1, showed approximately twofold upregulation of the exoS9-gfp reporter in comparison to PAO1. Upregulation of exoS9-gfp in the PDO100 mutant could be repressed to normal level by adding C4-HSL autoinducer, indicating a negative regulatory effect of RhlR/C4-HSL on exoS expression. As RhlR/C4-HSL is also involved in regulation of RpoS, the P. aeruginosa rpoS mutant SS24 was examined and the exoS9-gfp reporter was found to be fivefold upregulated in comparison to PAO1. For the first time evidence is reported for a regulatory cascade linking RhlR/RhlI and RpoS with the expression of the anti-host effector ExoS, part of the exoenzyme S regulon. Moreover, these data suggest that the exoenzyme S regulon may be downregulated in P. aeruginosa biofilms. INTRODUCTIONPseudomonas aeruginosa, an increasingly prevalent opportunistic human pathogen, is the most common Gram-negative bacterium found in infections of immunocompromised patients and of individuals suffering from cystic fibrosis (CF). Once P. aeruginosa has taken residence in the lungs of CF patients, it contributes substantially to morbidity and mortality in this population. The pseudomonads, which are seldom eradicated by antibacterial chemotherapy, chronically colonize the bronchoalveolar lumen of CF lungs and lead to recurrent pulmonal exacerbations, resulting in a progressive decline in lung function. Various virulence determinants have been shown to play a role in the pathogenesis of P. aeruginosa. The production of multiple of these virulence factors, including elastase, alkaline protease, LasA protease, phospholipase C, exotoxin A, rhamnolipid and pyocyanin relies on a cell-tocell communication system known as quorum sensing (van Delden & Iglewski, 1998;Winzer & Williams, 2001). These regulatory systems enable P. aeruginosa to produce virulence factors in a coordinated, cell-density-dependent manner. P. aeruginosa contains two separat...

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“…4,5 These proteins alter the function of the host cell, assist the microbe to invade, resist phagocytosis, grow in deep tissues and cause disease. [6][7][8][9][10] The T3SS is involved in a range of pathogenic mechanisms and its activity correlates closely with disease progression and outcome. 11 The well-studied plasmid-encoded Ysc of Yersinia is representative of these common virulence systems.…”

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confidence: 99%

Solving the Supply of Resveratrol Tetramers from Papua New Guinean Rainforest Anisoptera Species That Inhibit Bacterial Type III Secretion Systems

Davis

Beattie

et al. 2014

J. Nat. Prod.

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The supply of (−)-hopeaphenol (1) was achieved via enzymatic biotransformation in order to provide material for preclinical investigation. High-throughput screening of a prefractionated natural product library aimed to identify compounds that inhibit the bacterial virulence type III secretion system (T3SS) identified several fractions derived from two Papua New Guinean Anisoptera species, showing activity against Yersinia pseudotuberculosis outer proteins E and H (YopE and YopH). Bioassay-directed isolation from the leaves of A. thurifera, and similarly A. polyandra, resulted in three known resveratrol tetramers, (−)-hopeaphenol (1), vatalbinoside A (2), and vaticanol B (3). Compounds 1−3 displayed IC 50 values of 8.8, 12.5, and 9.9 μM in a luminescent reporter-gene assay (YopE) and IC 50 values of 2.9, 4.5, and 3.3 μM in an enzyme-based YopH assay, respectively, which suggested that they could potentially act against the T3SS in Yersinia. The structures of 1−3 were confirmed through a combination of spectrometric, chemical methods, and single-crystal X-ray structure determinations of the natural product 1 and the permethyl ether analogue of 3. The enzymatic hydrolysis of the β-glycoside 2 to the aglycone 1 was achieved through biotransformation using the endogenous leaf enzymes. This significantly enhanced the yield of the target bioactive natural product from 0.08% to 1.3% and facilitates ADMET studies of (−)-hopeaphenol (1).

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Triggering the ExoS regulon of Pseudomonas aeruginosa: A GFP-reporter analysis of exoenzyme (Exo) S, ExoT and ExoU synthesis

Cited by 48 publications

References 43 publications

Cheating by type 3 secretion system-negative Pseudomonas aeruginosa during pulmonary infection

Cheating by type 3 secretion system-negative Pseudomonas aeruginosa during pulmonary infection

Expression of Pseudomonas aeruginosa exoS is controlled by quorum sensing and RpoS

Solving the Supply of Resveratrol Tetramers from Papua New Guinean Rainforest Anisoptera Species That Inhibit Bacterial Type III Secretion Systems

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