Brefeldin A enhances Helicobacter pylori vacuolating cytotoxin-induced vacuolation of epithelial cells

Argent, Richard H.

doi:10.1016/j.femsle.2004.06.031

Cited by 6 publications

(6 citation statements)

References 17 publications

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“…However, we saw an increase in SRE-luciferase activity when cells were treated with BFA. Interestingly, VacA from Helicobacter pylori has also been shown to exhibit increased activity upon treatment with BFA [63]. There are several potential explanations for this increase.…”

Section: Discussionmentioning

confidence: 99%

Arf6-Dependent Intracellular Trafficking of Pasteurella multocida Toxin and pH-Dependent Translocation from Late Endosomes

Repella

Chong

et al. 2011

Toxins

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The potent mitogenic toxin from Pasteurella multocida (PMT) is the major virulence factor associated with a number of epizootic and zoonotic diseases caused by infection with this respiratory pathogen. PMT is a glutamine-specific protein deamidase that acts on its intracellular G-protein targets to increase intracellular calcium, cytoskeletal, and mitogenic signaling. PMT enters cells through receptor-mediated endocytosis and then translocates into the cytosol through a pH-dependent process that is inhibited by NH4Cl or bafilomycin A1. However, the detailed mechanisms that govern cellular entry, trafficking, and translocation of PMT remain unclear. Co-localization studies described herein revealed that while PMT shares an initial entry pathway with transferrin (Tfn) and cholera toxin (CT), the trafficking pathways of Tfn, CT, and PMT subsequently diverge, as Tfn is trafficked to recycling endosomes, CT is trafficked retrograde to the ER, and PMT is trafficked to late endosomes. Our studies implicate the small regulatory GTPase Arf6 in the endocytic trafficking of PMT. Translocation of PMT from the endocytic vesicle occurs through a pH-dependent process that is also dependent on both microtubule and actin dynamics, as evidenced by inhibition of PMT activity in our SRE-based reporter assay, with nocodazole and cytochalasin D, respectively, suggesting that membrane translocation and cytotoxicity of PMT is dependent on its transfer to late endosomal compartments. In contrast, disruption of Golgi-ER trafficking with brefeldin A increased PMT activity, suggesting that inhibiting PMT trafficking to non-productive compartments that do not lead to translocation, while promoting formation of an acidic tubulovesicle system more conducive to translocation, enhances PMT translocation and activity.

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Section: Discussionmentioning

confidence: 99%

Arf6-Dependent Intracellular Trafficking of Pasteurella multocida Toxin and pH-Dependent Translocation from Late Endosomes

Repella

Chong

et al. 2011

Toxins

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“…Vacuolation assay. AGS and MKN28 gastric epithelial cells were grown in 96-well plates until semi-confluent, before co-culture with H. pylori strains for 18-24 h (Argent et al, 2004a). After incubation, vacuolation was measured either by direct counting of randomly chosen microscopic fields or by use of a neutral red uptake assay (Cover et al, 1991).…”

Section: Methodsmentioning

confidence: 99%

Functional association between the Helicobacter pylori virulence factors VacA and CagA

Argent

Thomas

Letley

et al. 2008

Journal of Medical Microbiology

102

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The Helicobacter pylori virulence factors CagA and VacA are implicated in the development of gastroduodenal diseases. Most strains possessing CagA also possess the more virulent vacuolating form of VacA. This study assessed the significance of possession of both virulence factors in terms of their effect on gastric epithelial cells, using a set of minimally passaged, isogenic VacA, CagA and CagE mutants in H. pylori strains 60190 and 84-183. The cagA and cagE mutants were found to significantly increase VacA-induced vacuolation of epithelial cells, and the vacA mutants significantly increased CagA-induced cellular elongations, compared with wild-type strains, indicating that CagA reduces vacuolation and VacA reduces hummingbird formation. Although epithelial cells incubated with the wild-type H. pylori strains may display both vacuolation and hummingbird formation, it was found that (i) hummingbird length was significantly reduced in vacuolated cells compared with those without vacuolation; (ii) the number of vacuoles was significantly reduced in vacuolated cells with hummingbird formation compared with those without hummingbirds; and (iii) cells displaying extensive vacuolation did not subsequently form hummingbirds and vice versa. VacA did not affect the phosphorylation of CagA. These data show that VacA and CagA downregulate each other's effects on epithelial cells, potentially allowing H. pylori interaction with cells whilst avoiding excessive cellular damage. INTRODUCTIONHelicobacter pylori strains infect the stomachs of half of the world's population. They cause gastritis and gastric and duodenal ulceration, and are a major risk factor for the development of mucosa-associated lymphoid tissue lymphoma and gastric adenocarcinoma. A number of virulence factors are associated with disease outcome, including the vacuolating cytotoxin (VacA) and the possession of the cytotoxin-associated gene pathogenicity island (cag PAI) comprising 27-31 genes (Censini et al., 1996).The vacA gene is present in virtually all strains of H. pylori but is polymorphic (Atherton et al., 1997), comprising variable signal regions (type s1 or s2) and mid-regions (type m1 or m2). Type s1/m1 VacA causes more epithelial cell damage than type s1/m2, whereas type s2/m2 and the rare s2/m1 are non-toxic due to the presence of a short 12-residue hydrophilic extension on the s2 form . VacA forms anion-selective channels within artificial membranes (Czajkowsky et al., 1999) and is assumed to do the same in vivo, increasing permeability to anions and urea (Tombola et al., 2001). Endocytosis of VacA channels leads to the formation of large vacuoles within the late endosome-lysosome compartment.The cag PAI encodes a type IV secretory system that causes inflammation by activation of NF-kB and secretion of cytokines and chemokines such as interleukin 8 (IL-8) Censini et al., 1996;Keates et al., 1997;Viala et al., 2004;Brandt et al., 2005), and facilitates the translocation of CagA into the cytosol of epithelial cells, where it becomes tyrosine phosp...

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“…VacA-induced vacuolation of AGS and RK13 epithelial cells was determined microscopically after coculture with H. pylori strains for 16 to 24 h, as described (40).…”

Section: Methodsmentioning

confidence: 99%

Toxigenic Helicobacter pylori Infection Precedes Gastric Hypochlorhydria in Cancer Relatives, and H. pylori Virulence Evolves in These Families

Argent

Thomas

Avilés-Jiménez

et al. 2008

Clinical Cancer Research

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Purpose: Helicobacter pylori infection by virulent strains is associated with gastric adenocarcinoma. We aimed to determine whether infection with virulent H. pylori preceeded precancerous gastric hypochlorhydria and atrophy in gastric cancer relatives and quantify the extent of virulence factor evolution. Experimental Design: H. pylori strains from 51Scottish gastric cancer relatives were characterized by genetic fingerprinting and typing the vacuolating cytotoxin gene (vacA), the cytotoxinassociated gene (cagA), and housekeeping genes. We phenotyped strains by coculture with gastric epithelial cells and assessing vacuolation (microscopy), CagA tyrosine phosphorylation (immunoblot), and interleukin-8 secretion (ELISA). Results: Toxigenic (vacA type s1/m1) H. pylori was associated with precancerous gastric hypochlorhydria (P < 0.01). Adult family members with this type of H. pylori had the same strain as currently noncohabiting adult family members in 68 % cases, implying acquisition during childhood from each other or a common source.We analyzed different isolates of the same strain within families and showed that H. pylori commonly microevolved to change virulence: this occurred in 22% individuals and a striking 44% cases where the strain was shared within families. Microevolution in vacA occurred by extragenomic recombination and in cagA by this or duplication/deletion. Microevolution led to phenotypic changes in virulence. Passage of microevolved strains could be tracked within families. Conclusions:Toxigenic H. pylori infection precedes and so likely causes gastric hypochlorhydria, suggesting that virulent H. pylori increases cancer risk by causing this condition. Microevolution of virulence genes is common within families of gastric cancer patients and changes H. pylori virulence.

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Brefeldin A enhances Helicobacter pylori vacuolating cytotoxin-induced vacuolation of epithelial cells

Cited by 6 publications

References 17 publications

Arf6-Dependent Intracellular Trafficking of Pasteurella multocida Toxin and pH-Dependent Translocation from Late Endosomes

Arf6-Dependent Intracellular Trafficking of Pasteurella multocida Toxin and pH-Dependent Translocation from Late Endosomes

Functional association between the Helicobacter pylori virulence factors VacA and CagA

Toxigenic Helicobacter pylori Infection Precedes Gastric Hypochlorhydria in Cancer Relatives, and H. pylori Virulence Evolves in These Families

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