A Type III Secretion Negative Clinical Strain of Pseudomonas aeruginosa Employs a Two-Partner Secreted Exolysin to Induce Hemorrhagic Pneumonia

Elsen, Sylvie; Huber, Philippe; Bouillot, Stéphanie; Couté, Yohann; Fournier, P; Dubois, Yohann; Timsit, Jean‐François; Maurin, Max; Attrée, Ina

doi:10.1016/j.chom.2014.01.003

Cited by 85 publications

(155 citation statements)

References 59 publications

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“…One of these contains strains previously referred to as “clonal outliers” (5–7); the first identified member of this group is strain PA7 (8). Although the virulence of P. aeruginosa is multifactorial, the two clades differ mainly in the way that they exert their cytolytic activity on human cells (6, 9, 10).…”

Section: Introductionmentioning

confidence: 99%

Pseudomonas aeruginosa Pore-Forming Exolysin and Type IV Pili Cooperate To Induce Host Cell Lysis

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Elsen

et al. 2017

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Clinical strains of Pseudomonas aeruginosa lacking the type III secretion system genes employ a toxin, exolysin (ExlA), for host cell membrane disruption. Here, we demonstrated that ExlA export requires a predicted outer membrane protein, ExlB, showing that ExlA and ExlB define a new active two-partner secretion (TPS) system of P. aeruginosa. In addition to the TPS signals, ExlA harbors several distinct domains, which include one hemagglutinin domain, five arginine-glycine-aspartic acid (RGD) motifs, and a C-terminal region lacking any identifiable sequence motifs. However, this C-terminal region is important for the toxic activity, since its deletion abolishes host cell lysis. Using lipid vesicles and eukaryotic cells, including red blood cells, we demonstrated that ExlA has a pore-forming activity which precedes cell membrane disruption of nucleated cells. Finally, we developed a high-throughput cell-based live-dead assay and used it to screen a transposon mutant library of an ExlA-producing P. aeruginosa clinical strain for bacterial factors required for ExlA-mediated toxicity. The screen resulted in the identification of proteins involved in the formation of type IV pili as being required for ExlA to exert its cytotoxic activity by promoting close contact between bacteria and the host cell. These findings represent the first example of cooperation between a pore-forming toxin of the TPS family and surface appendages in host cell intoxication.

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Section: Introductionmentioning

confidence: 99%

Pseudomonas aeruginosa Pore-Forming Exolysin and Type IV Pili Cooperate To Induce Host Cell Lysis

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Ragno

Elsen

et al. 2017

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“…Cell death was evaluated by measuring the lactate dehydrogenase (LDH) released in the supernatant, as previously described (21). Briefly, strains were grown to late exponential phase (A 600 ϭ 1) and used to infect cells at an MOI of 10.…”

Section: Methodsmentioning

confidence: 99%

Pharmacological Activation of Rap1 Antagonizes the Endothelial Barrier Disruption Induced by Exotoxins ExoS and ExoT of Pseudomonas aeruginosa

2015

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Most clinical strains of Pseudomonas aeruginosa, a leading agent of nosocomial infections, are multiresistant to antibiotherapy. Because of the paucity of new available antibiotics, the investigation of strategies aimed at limiting the action of its major virulence factors has gained much interest. The type 3 secretion system of P. aeruginosa and its effectors are known to be major determinants of toxicity and are required for bacterial dissemination in the host. Bacterial transmigration across the vascular wall is considered to be an important step in the infectious process. Using human endothelial primary cells, we demonstrate that forskolin (FSK), a drug inducing cyclic AMP (cAMP) elevation in eukaryotic cells, strikingly reduced the cell retraction provoked by two type 3 toxins, ExoS and ExoT, found in the majority of clinical strains. Conversely, cytotoxicity of a strain carrying the type 3 effector ExoU was unaffected by FSK. In addition, FSK altered the capacity of two ExoS/ExoT strains to transmigrate across cell monolayers. In agreement with these findings, other drugs and a cytokine inducing the increase of cAMP intracellular levels have also protected cells from retraction. cAMP is an activator of both protein kinase A and EPAC, a GTPase exchange factor of Rap1. Using activators or inhibitors of either pathway, we show that the beneficial effect of FSK is exerted by the activation of the EPAC/Rap1 axis, suggesting that its protective effect is mediated by reinforcing cell-cell and cell-substrate adhesion. Pseudomonas aeruginosa is an opportunistic pathogen and a leading agent of nosocomial infections. The largest cohorts of P. aeruginosa-infected patients are found in three pathological settings: ventilator-associated pneumonia, bacteremia, and cystic fibrosis. In acute infections, P. aeruginosa disseminates from the primary infection site to the blood and other organs, leading to sepsis and multiple organ failure. From a clinical point of view, vascular barrier breakdown is thus considered to be a key step in the pathophysiology of infection (1).Most P. aeruginosa clinical isolates are multidrug or even extremely drug resistant to antibiotics, which explains the high fatality rates of P. aeruginosa infections. The pathogen has been recently included in a family of so-called "ESKAPE" bacterial pathogens, a group which also includes Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, and Enterobacter species, that are able to efficiently "escape" the effects of available antibacterial drugs and for which there is an urgent need for developments of novel types of drugs (2, 3). In this context, the investigation of new strategies limiting the action of the virulence factors rather than bactericidal agents has gained much interest.P. aeruginosa is extremely well equipped in virulence determinants, which are membrane-embedded protein machineries dedicated for effector/toxin export (4). The type 3 secretion system (T3SS) and its effectors are recognized as the most i...

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“…Elsen and colleagues identified another member of the PA7 family, CLJ1, that was cultured from a patient with severe hemorrhagic pneumonia (9). This prompted them to ask the question of how this strain could be so virulent in the absence of type 3 secretion.…”

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“…This prompted them to ask the question of how this strain could be so virulent in the absence of type 3 secretion. In their recent publication, they describe a novel and very interesting cytotoxin produced by CLJ1 (9). This toxin shares homology with hemolysins found in several Gram-negative pathogens and was termed Exolysin.…”

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Different paths to pathogenesis

Allen

Ozer

Hauser

2014

Trends in Microbiology

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A Type III Secretion Negative Clinical Strain of Pseudomonas aeruginosa Employs a Two-Partner Secreted Exolysin to Induce Hemorrhagic Pneumonia

Cited by 85 publications

References 59 publications

Pseudomonas aeruginosa Pore-Forming Exolysin and Type IV Pili Cooperate To Induce Host Cell Lysis

Pseudomonas aeruginosa Pore-Forming Exolysin and Type IV Pili Cooperate To Induce Host Cell Lysis

Pharmacological Activation of Rap1 Antagonizes the Endothelial Barrier Disruption Induced by Exotoxins ExoS and ExoT of Pseudomonas aeruginosa

Different paths to pathogenesis

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