Alteration of pulmonary structure by Pseudomonas aeruginosa exoenzyme S

Woods, Donald E.; Hwang, W. S.; Shahrabadi, M Shamsi; Que, Johnu .

doi:10.1099/00222615-26-2-133

Cited by 37 publications

(23 citation statements)

References 20 publications

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“…The vacuoles induced by the cytotoxin in vitro were, therefore, characterised cytochemically and ultrastructurally to understand better the mechanism of action of the cytotoxin and its potential clinical relevance. Vacuolation of mammalian cells have been reported to follow nutrient deprivation, chemical exposure and bacterial toxin activity.12* 239 24 One such vacuole type, the autophagosome, normally contributes to the recycling of cytoplasmic components, but appears rapidly and in excessive numbers in cells exposed to metabolic stress12* 13* l7 and is believed to originate by fusion of ER-derived vacuoles and lysosomes.12T l3 Autophagosomes are acidic and contain acid phosphatase activity and cytoplasmic debris. The present study shows that toxin-induced vacuoles resemble autophagosomes in the nature of their contents and their communication with the endo-lysosomal pathway during maturation.…”

Section: Discussionmentioning

confidence: 99%

Character and origin of vacuoles induced in mammalian cells by the cytotoxin of Helicobacter pylori

Catrenich

Chestnut

1992

Journal of Medical Microbiology

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Summary. Cytotoxic activity of culture supernates of Helicobacter pylori is manifested by vacuolation of mammalian cells in vitro. The formation and maturation of toxin-induced vacuoles in HeLa cells has been studied to examine the possibility that they are autophagosomal in nature. Observation by light microscopy revealed that vacuoles originate in a perinuclear position, increasing in number and size until cell degeneration and lysis occur after 48 h. Ultrastructural study of mature vacuoles indicated the presence of a bounding membrane with contents consisting of degenerate cytoplasmic components and acid phosphatase activity. Confocal fluorescence imaging demonstrated acridine orange accumulation in the vacuoles of toxin-treated cells, indicating an acidic intravacuolar pH. These features are characteristic of autophagosomes. In addition, the size of vacuoles in living, acridine orange-stained cells tended to be inversely proportional to fluorescence intensity. Fluid phase endocytic markers were observed only rarely within nascent vacuoles. Over the succeeding 24 h, labelling of most vacuoles with these dyes was observed. This, along with the observation of intravacuolar acid phosphatase activity, provides evidence that vacuoles communicate at some point during their development with endocytically derived compartments. These observations provide direct evidence for an autophagic origin of these structures.

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Section: Discussionmentioning

confidence: 99%

Character and origin of vacuoles induced in mammalian cells by the cytotoxin of Helicobacter pylori

Catrenich

Chestnut

1992

Journal of Medical Microbiology

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“…Prototrophic strains of P. aeruginosa used in this study include PAO1 (14), DG1 (34), FRD2, a spontaneous nonmucoid revertant of the cystic fibrosis isolate FRD1 (8), PA103, an elastase-negative strain (25), and PAO-E64, a lasAl mutant of PAO1 which is elastolytic deficient (24).…”

Section: Methodsmentioning

confidence: 99%

Efficient production and processing of elastase and LasA by Pseudomonas aeruginosa require zinc and calcium ions

Olson

Ohman

1992

J Bacteriol

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The ability of Pseudomonas aeruginosa to degrade elastin, a major component of connective tissue, likely contributes to its pathogenicity and multiplication in human tissues. Two extracellular enzymes are required for P. aeruginosa elastolytic activity: elastase and LasA. Elastase is a zinc metalloprotease, but little is known about the structure of LasA. When grown under metal ion-deficient conditions, P. aeruginosa culture supernatants were found to exhibit a low level of elastolytic activity, which coincided with production of low levels of the 51-kDa proelastase and no detectable LasA. By using this fact to identify factors that promote elastolytic activity, P. aeruginosa PAO1, FRD2, and DG1 were grown in metal ion-deficient medium supplemented with zinc (l0-4 M ZnCl2), calcium (2.5 x i0-3 M CaCl2), or iron (1O-4 M FeCl3). High levels of proteolytic and elastolytic activity were exhibited by all strains when cultured in the presence of both zinc and calcium, and this was associated with the production of mature 33-kDa elastase and 21-kDa LasA. Supplementing DG1 and PAO1 cultures with zinc alone stimulated the production of 33-kDa elastase, which, because of the calcium-deficient conditions, exhibited low proteolytic and elastolytic activities. Zinc also stimulated the production of a 41-kDa form of LasA in DG1 and PAO1 culture supernatants. Elastase production by FRD2 cultured in the presence of zinc alone differed from that by the other two strains in that supernatants contained 33-kDa elastase, a 21-kDa form of LasA, and exhibited high proteolytic and elastolytic activities. Such strain-associated differences in LasA processing and elastase activity can be explained by differences in metal ion-scavenging mechanisms adapted by the strains. Supplementing cultures with calcium stimulated the production of elastase but had no effect on LasA production. The elastase produced exhibited variable sizes, possibly resulting from aberrant processing reactions, and showed little proteolytic activity. Proteolytic activity could be recovered from 33-kDa elastase produced in the presence of calcium by inclusion of zinc in the enzymatic assay. Although iron was previously found to exert a repressive effect on P. aeruginosa elastolytic activity, iron exerted little effect on elastolytic activity when added to cultures containing both zinc and calcium. These studies support the conclusion that elastase production and processing are promoted by both zinc and calcium. LasA production, in comparison, is stimulated by zinc, with both zinc and calcium facilitating its processing. The association of 41-kDa LasA with a low level of elastolytic activity and of 21-kDa LasA with a high level of activity supports the conclusion that lasA encodes a larger, precursor protein which is processed to an active 21-kDa form during secretion.Pseudomonas aeruginosa is an opportunistic pathogen known to cause severe and lethal infections in compromised hosts. Elastase is one of several extracellular enzymes produced by P. aeruginosa that is beli...

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“…One of the hallmarks of the lung damage in CF is an ineffective inflammatory response that results in severe neutrophil-mediated pulmonary damage and an inability to clear the organisms. P. aeruginosa contributes to the lung damage by the production of virulence factors; one of the most important virulence factors, exoenzyme S, has been shown to induce pulmonary damage in animal models (17,29,35), and increased levels of exoenzyme S correlate with human disease (18,36). The cytotoxicity of exoenzyme S for epithelial cells follows both contact-dependent type III translocation into eukaryotic target cells and contact-independent type III secretion, suggesting two mechanisms of cellular activation: intracellular and extracellular (15,34,35).…”

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confidence: 99%

“…ExoS/DG1 from P. aeruginosa strain DG1 was isolated as previously described using (NH 4 ) 2 SO 4 precipitation of culture supernatants, ion-exchange chromatography, and acetone precipitation, followed by gel filtration, and migrated as a 50-kDa band without ADP-ribosyltransferase activity (35). rExoS was isolated from Escherichia coli BL21(DE3) bearing a plasmid encoding histidine-tagged exoenzyme S cloned from P. aeruginosa 388(pETrHisExoS).…”

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confidence: 99%

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Pseudomonas aeruginosaExoenzyme S Induces Transcriptional Expression of Proinflammatory Cytokines and Chemokines

Epelman

Bruno

Neely³

et al. 2000

Infect Immun

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Pseudomonas aeruginosa infection of cystic fibrosis patients causes lung damage that is substantially orchestrated by cytokines. In this study, multi-gene probe analysis was used to characterize the ability of the P. aeruginosa mitogen, exoenzyme S, to induce proinflammatory and immunoregulatory cytokines and chemokines. Exoenzyme S strongly induced transcription of proinflammatory cytokines and chemokines (tumor necrosis factor alpha, interleukin-1␣ [IL-1␣], IL-1␤, IL-6, IL-8, MIP-1␣, MIP-1␤, MCP-1, RANTES, and I-309), modest transcription of immunoregulatory cytokines (IL-10 and IL-12p40), and weak transcription of Th1 cytokines (IL-2 and gamma interferon). The response occurred early and subsided without evolving over time. These data suggest that cells responding to exoenzyme S would rapidly express proinflammatory cytokines and chemokines that may contribute to pulmonary inflammation in cystic fibrosis.Virtually all cystic fibrosis (CF) patients are colonized by Pseudomonas aeruginosa, and 90% of those that are colonized die as a result of lung damage (11). One of the hallmarks of the lung damage in CF is an ineffective inflammatory response that results in severe neutrophil-mediated pulmonary damage and an inability to clear the organisms. P. aeruginosa contributes to the lung damage by the production of virulence factors; one of the most important virulence factors, exoenzyme S, has been shown to induce pulmonary damage in animal models (17,29,35), and increased levels of exoenzyme S correlate with human disease (18, 36). The cytotoxicity of exoenzyme S for epithelial cells follows both contact-dependent type III translocation into eukaryotic target cells and contact-independent type III secretion, suggesting two mechanisms of cellular activation: intracellular and extracellular (15,34,35). We have recently described an additional activity: extracellular exoenzyme S is mitogenic for T cells, inducing tremendous T-cell activation (5-7). Exoenzyme-S-induced activation of T cells is neither dependent upon nor inhibited by ADP-ribosyltransferase activity, and this activity is present in exoenzyme S from both purified and recombinant sources (7). Further, we have found that activation by exoenzyme S induces T-cell apoptosis (6). The current studies were performed to determine whether this T-cell activation culminates in the induction of cytokines that have the potential of influencing immunoinflammatory responses or whether apoptosis precludes cytokine transcription.Immunoinflammatory responses are orchestrated by cytokines, and T-cell mitogens and superantigens are potent stimuli for cytokine production from T cells, monocytes, and macrophages (2, 3). In CF there is a misdirected or dysregulated response, since there is a chronic and exuberant immunoinflammatory response without clearance of the pathogen. This response may be a result of altered cytokine induction that includes decreased secretion of the anti-inflammatory cytokine interleukin-10 (IL-10) (13, 28) with concordant increases in proinflammatory cy...

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Alteration of pulmonary structure by Pseudomonas aeruginosa exoenzyme S

Cited by 37 publications

References 20 publications

Character and origin of vacuoles induced in mammalian cells by the cytotoxin of Helicobacter pylori

Character and origin of vacuoles induced in mammalian cells by the cytotoxin of Helicobacter pylori

Efficient production and processing of elastase and LasA by Pseudomonas aeruginosa require zinc and calcium ions

Pseudomonas aeruginosaExoenzyme S Induces Transcriptional Expression of Proinflammatory Cytokines and Chemokines

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