Browse
Submit Data
Databases
API
Help

Dataset Information

38 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Outer membrane targeting of passenger proteins by the vacuolating cytotoxin autotransporter of Helicobacter pylori.

ABSTRACT: Helicobacter pylori produces a number of proteins associated with the outer membrane, including adhesins and the vacuolating cytotoxin. These proteins are supposed to integrate into the outer membrane by beta-barrel structures, characteristic of the family of autotransporter proteins. By using the SOMPES (shuttle vector-based outer membrane protein expression) system for outer membrane protein production, we were able to functionally express in H. pylori the cholera toxin B subunit genetically fused to the C-terminal VacA domain. We demonstrate that the fusion protein is translocated to the H. pylori outer membrane and that the CtxB domain is exposed on the H. pylori surface. Thus, we provide the first experimental evidence that the C-terminal beta-domain of VacA can transport a foreign passenger protein to the H. pylori surface and hence acts as a functional autotransporter.

SUBMITTER: Fischer W

PROVIDER: S-EPMC100054 | biostudies-literature | 2001 Nov

REPOSITORIES: biostudies-literature

ACCESS DATA

Json Xml

Publications

Outer membrane targeting of passenger proteins by the vacuolating cytotoxin autotransporter of Helicobacter pylori.

Fischer W W Buhrdorf R R Gerland E E Haas R R

Infection and immunity 20011101 11

Helicobacter pylori produces a number of proteins associated with the outer membrane, including adhesins and the vacuolating cytotoxin. These proteins are supposed to integrate into the outer membrane by beta-barrel structures, characteristic of the family of autotransporter proteins. By using the SOMPES (shuttle vector-based outer membrane protein expression) system for outer membrane protein production, we were able to functionally express in H. pylori the cholera toxin B subunit genetically f ...[more]

PMID: 11598049

Similar Datasets

Evolution of the Helicobacter pylori vacuolating cytotoxin in a human stomach.

Project description:We describe two subclones of Helicobacter pylori, isolated contemporaneously from a human stomach, which differ markedly in the vacuolating cytotoxin gene, vacA, but whose near identity in sequences outside this locus implies a very recent common origin. The differences are consistent with homologous recombination with DNA from another strain and result in a changed vacA midregion and, importantly, in changed toxicity.

| S-EPMC451639 | biostudies-literature

DNA sequence analysis of South African Helicobacter pylori Vacuolating Cytotoxin Gene (vacA).

Project description:Sequence diversity and population structures can vary widely among pathogenic bacteria species. In some species, all isolates are highly similar, whereas in others most of the isolates are distinguished easily. H. pylori is known for its wide genetic diversity amongst the various strains most especially in the genes involved in virulence. The aim of this study was to evaluate by PCR and sequence analysis, the genetic profile of H. pylori vacA gene (s1, s2, m1 and m2). We sequenced small DNA segments from 13 vacAs1, 10 vacAm2, 6 vacAm1 and 6 vacAs2 strains which were amplified with amplicon size of 259/286 bp, 290 bp and 352 bp for vacAs1/s2, m1 and m2 respectively. Based on similarities among our strains accession numbers were provided for seven vacAs1 (HQ709109-HQ709115), six vacAs2 (JN848463-JN848468), six vacAm1 (JN848469-JN848474) and six vacAm2 (HQ650801-HQ650806) strains. Amongst the strains studied, 98.07%, 98.58%, 97.38% and 95.41% of vacAs1, vacAs2, vacAm1 and vacAm2 of the strains were conserved respectively. Findings of this study underscores the importance of understanding the virulence composition and diversity of H. pylori in South Africa for enhanced clinico-epidemiological monitoring and pathophysiology of disease.

| S-EPMC3233416 | biostudies-literature

Cryo-EM structures of Helicobacter pylori vacuolating cytotoxin A (VacA) oligomeric assemblies

Project description: Not available

| EMPIAR-10607 | biostudies-other

Natural diversity in the N terminus of the mature vacuolating cytotoxin of Helicobacter pylori determines cytotoxin activity.

Project description:Naturally occurring noncytotoxic vacA type s2 strains of Helicobacter pylori have a 12-residue extension to the vacuolating cytotoxin (VacA) compared with cytotoxic type s1 strains. We show that adding the region encoding this extension to type s1 vacA completely abolishes vacuolating cytotoxin activity but has no effect on VacA production.

| S-EPMC94518 | biostudies-literature

Heterogeneity in levels of vacuolating cytotoxin gene (vacA) transcription among Helicobacter pylori strains.

Project description:Broth culture supernatants from Tox+ Helicobacter pylori strains induce vacuolation of HeLa cells in vitro and contain VacA in concentrations that are higher than those found in supernatants from Tox- H. pylori strains. To investigate the basis for this phenomenon, we analyzed the transcription of the vacuolating cytotoxin gene (vacA) in eight Tox+ strains (each with a type s1/m1 vacA genotype) and nine Tox- strains (each with a type s2/m2 vacA genotype). Most of the Tox+ and Tox- strains tested used the same vacA transcriptional start point, but Tox+ strains yielded significantly stronger primer extension signal intensities than did Tox- strains (mean densitometry values of 15.8 +/- 1.9 versus 8.9 +/- 1.7, P = 0. 0016). Correspondingly, when we introduced vacA::xylE transcriptional fusions into the chromosomes of a Tox+ strain (60190) and a Tox- strain (86-313), the level of XylE activity in 60190 vacA::xylE was about 30-fold higher than that in 86-313 vacA::xylE. Sequence analysis and promoter exchange experiments indicated that the different levels of vacA transcription in these two strains cannot be explained solely by a difference in promoter strength. These data indicate that Tox+ and Tox- H. pylori strains typically differ not only in the VacA amino acid sequence but also in the level of vacA transcription.

| S-EPMC108317 | biostudies-literature

Helicobacter pylori γ-glutamyl transpeptidase and vacuolating cytotoxin promote gastric persistence and immune tolerance.

Project description:Infection with the gastric bacterial pathogen Helicobacter pylori is typically contracted in early childhood and often persists for decades. The immunomodulatory properties of H. pylori that allow it to colonize humans persistently are believed to also account for H. pylori's protective effects against allergic and chronic inflammatory diseases. H. pylori infection efficiently reprograms dendritic cells (DCs) toward a tolerogenic phenotype and induces regulatory T cells (Tregs) with highly suppressive activity in models of allergen-induced asthma. We show here that two H. pylori virulence determinants, the γ-glutamyl transpeptidase GGT and the vacuolating cytotoxin VacA, contribute critically and nonredundantly to H. pylori's tolerizing effects on murine DCs in vitro and in vivo. The tolerance-promoting effects of both factors are independent of their described suppressive activity on T cells. Isogenic H. pylori mutants lacking either GGT or VacA are incapable of preventing LPS-induced DC maturation and fail to drive DC tolerization as assessed by induction of Treg properties in cocultured naive T cells. The Δggt and ΔvacA mutants colonize mice at significantly reduced levels, induce stronger T-helper 1 (Th1) and T-helper 17 (Th17) responses, and/or trigger more severe gastric pathology. Both factors promote the efficient induction of Tregs in vivo, and VacA is required to prevent allergen-induced asthma. The defects of the Δggt mutant in vitro and in vivo are phenocopied by pharmacological inhibition of the transpeptidase activity of GGT in all readouts. In conclusion, our results reveal the molecular players and mechanistic basis for H. pylori-induced immunomodulation, promoting persistent infection and conferring protection against allergic asthma.

| S-EPMC3581963 | biostudies-literature

The m2 form of the Helicobacter pylori cytotoxin has cell type-specific vacuolating activity.

Project description:The Helicobacter pylori toxin VacA causes vacuolar degeneration in mammalian cell lines in vitro and plays a key role in peptic ulcer disease. Two alleles, m1 and m2, of the mid-region of the vacA gene have been described, and the m2 cytotoxin always has been described as inactive in the in vitro HeLa cell assay. However, the m2 allele is associated with peptic ulcer and is prevalent in populations in which peptic ulcer and gastric cancer have high incidence. In this paper, we show that, despite the absence of toxicity on HeLa cells, the m2 cytotoxin is able to induce vacuolization in primary gastric cells and in other cell lines such as RK-13. The absence of Hela cell activity is due to an inability to interact with the cell surface, suggesting a receptor-mediated interaction. This result is consistent with the observation that the m2 allele is found in a population that has a high prevalence of peptic ulcer disease and gastric cancer. VacA is the first bacterial toxin described for which the same active subunit can be delivered by different receptor binding domains.

| S-EPMC21487 | biostudies-literature

Amino-terminal hydrophobic region of Helicobacter pylori vacuolating cytotoxin (VacA) mediates transmembrane protein dimerization.

Project description:Helicobacter pylori VacA is a secreted protein toxin that forms channels in lipid bilayers and induces multiple structural and functional alterations in eukaryotic cells. A unique hydrophobic segment at the amino terminus of VacA contains three tandem repeats of a GxxxG motif that is characteristic of transmembrane dimerization sequences. To examine functional properties of this region, we expressed and analyzed ToxR-VacA-maltose binding protein fusions using the TOXCAT system, which was recently developed by W. P. Russ and D. M. Engelman (Proc. Natl. Acad. Sci. USA 96:863-868, 1999) to study transmembrane helix-helix associations in a natural membrane environment. A wild-type VacA hydrophobic region mediated insertion of the fusion protein into the inner membrane of Escherichia coli and mediated protein dimerization. A fusion protein containing a mutant VacA hydrophobic region (in which glycine 14 of VacA was replaced by alanine) also inserted into the inner membrane but dimerized significantly less efficiently than the fusion protein containing the wild-type VacA sequence. Based on these results, we speculate that the wild-type VacA amino-terminal hydrophobic region contributes to oligomerization of the toxin within membranes of eukaryotic cells.

| S-EPMC98002 | biostudies-literature

Vacuolating cytotoxin and variants in Atg16L1 that disrupt autophagy promote Helicobacter pylori infection in humans.

Project description:The Helicobacter pylori toxin vacuolating cytotoxin (VacA) promotes gastric colonization, and its presence (VacA(+)) is associated with more-severe disease. The exact mechanisms by which VacA contributes to infection are unclear. We previously found that limited exposure to VacA induces autophagy of gastric cells, which eliminates the toxin; we investigated whether autophagy serves as a defense mechanism against H pylori infection.We investigated the effect of VacA on autophagy in human gastric epithelial cells and primary gastric cells from mice. Expression of p62, a marker of autophagy, was also assessed in gastric tissues from patients infected with toxigenic (VacA(+)) or nontoxigenic strains. We analyzed the effect of VacA on autophagy in peripheral blood monocytes obtained from subjects with different genotypes of ATG16L1, which regulates autophagy. We performed genotyping for ATG16L1 in 2 cohorts of infected and uninfected subjects.Prolonged exposure of human gastric epithelial cells and mouse gastric cells to VacA disrupted induction of autophagy in response to the toxin, because the cells lacked cathepsin D in autophagosomes. Loss of autophagy resulted in the accumulation of p62 and reactive oxygen species. Gastric biopsy samples from patients infected with VacA(+), but not nontoxigenic strains of H pylori, had increased levels of p62. Peripheral blood monocytes isolated from individuals with polymorphisms in ATG16L1 that increase susceptibility to Crohn's disease had reduced induction of autophagy in response to VacA(+) compared to cells from individuals that did not have these polymorphisms. The presence of the ATG16L1 Crohn's disease risk variant increased susceptibility to H pylori infection in 2 separate cohorts.Autophagy protects against infection with H pylori; the toxin VacA disrupts autophagy to promote infection, which could contribute to inflammation and eventual carcinogenesis.

| S-EPMC3336037 | biostudies-literature

Cryo-EM structures of Helicobacter pylori vacuolating cytotoxin A oligomeric assemblies at near-atomic resolution.

Project description:Human gastric pathogen Helicobacter pylori (H. pylori) is the primary risk factor for gastric cancer and is one of the most prevalent carcinogenic infectious agents. Vacuolating cytotoxin A (VacA) is a key virulence factor secreted by H. pylori and induces multiple cellular responses. Although structural and functional studies of VacA have been extensively performed, the high-resolution structure of a full-length VacA protomer and the molecular basis of its oligomerization are still unknown. Here, we use cryoelectron microscopy to resolve 10 structures of VacA assemblies, including monolayer (hexamer and heptamer) and bilayer (dodecamer, tridecamer, and tetradecamer) oligomers. The models of the 88-kDa full-length VacA protomer derived from the near-atomic resolution maps are highly conserved among different oligomers and show a continuous right-handed ?-helix made up of two domains with extensive domain-domain interactions. The specific interactions between adjacent protomers in the same layer stabilizing the oligomers are well resolved. For double-layer oligomers, we found short- and/or long-range hydrophobic interactions between protomers across the two layers. Our structures and other previous observations lead to a mechanistic model wherein VacA hexamer would correspond to the prepore-forming state, and the N-terminal region of VacA responsible for the membrane insertion would undergo a large conformational change to bring the hydrophobic transmembrane region to the center of the oligomer for the membrane channel formation.

| S-EPMC6452728 | biostudies-literature

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

Tweets

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data