title: Characterization of Helicobacter pylori VacA-containing vacuoles (VCVs), VacA intracellular trafficking and interference with calcium signalling in T lymphocytes.
abstract: The human pathogen Helicobacter pylori colonizes half of the global population. Residing at the stomach epithelium, it contributes to the development of diseases such as gastritis, duodenal and gastric ulcers, and gastric cancer. A major factor is the secreted vacuolating toxin VacA, which forms anion-selective channels in the endosome membrane that cause the compartment to swell, but the composition and purpose of the resulting VacA-containing vacuoles (VCVs) are still unknown. VacA exerts influence on the host immune response in various ways, including inhibition of T-cell activation and proliferation and suppression of the host immune response. In this study, for the first time the composition of VCVs from T cells was comprehensively analysed to investigate VCV function. VCVs were successfully isolated via immunomagnetic separation, and the purified vacuoles were analysed by mass spectrometry. We detected a set of 122 VCV-specific proteins implicated among others in immune response, cell death and cellular signalling processes, all of which VacA is known to influence. One of the individual proteins studied further was stromal interaction molecule (STIM1), a calcium sensor residing in the endoplasmic reticulum (ER) that is important in store-operated calcium entry. Live cell imaging microscopy data demonstrated colocalization of VacA with STIM1 in the ER and indicated that VacA may interfere with the movement of STIM1 towards the plasma membrane-localized calcium release activated calcium channel protein ORAI1 in response to Ca(2+) store depletion. Furthermore, VacA inhibited the increase of cytosolic-free Ca(2+) in the Jurkat E6-1 T-cell line and human CD4(+) T cells. The presence of VacA in the ER and its trafficking to the Golgi apparatus was confirmed in HeLa cells, identifying these two cellular compartments as novel VacA target structures.
authors.affiliation: Max von Pettenkofer-Institut für Hygiene und Medizinische Mikrobiologie, Ludwig-Maximilians-Universität, München, Germany.
authors.forename: Beate
authors.initials: B
authors.lastname: Kern
authors.affiliation: Max von Pettenkofer-Institut für Hygiene und Medizinische Mikrobiologie, Ludwig-Maximilians-Universität, München, Germany.
authors.forename: Utkarsh
authors.initials: U
authors.lastname: Jain
authors.affiliation: Max von Pettenkofer-Institut für Hygiene und Medizinische Mikrobiologie, Ludwig-Maximilians-Universität, München, Germany.
authors.forename: Ciara
authors.initials: C
authors.lastname: Utsch
authors.affiliation: Institut für Mikrobiologie, Ernst-Moritz-Arndt Universität Greifswald, Greifswald, Germany.
authors.forename: Andreas
authors.initials: A
authors.lastname: Otto
authors.affiliation: Max von Pettenkofer-Institut für Hygiene und Medizinische Mikrobiologie, Ludwig-Maximilians-Universität, München, Germany.
authors.forename: Benjamin
authors.initials: B
authors.lastname: Busch
authors.affiliation: Max von Pettenkofer-Institut für Hygiene und Medizinische Mikrobiologie, Ludwig-Maximilians-Universität, München, Germany.
authors.forename: Luisa
authors.initials: L
authors.lastname: Jiménez-Soto
authors.affiliation: Institut für Mikrobiologie, Ernst-Moritz-Arndt Universität Greifswald, Greifswald, Germany.
authors.forename: Dörte
authors.initials: D
authors.lastname: Becher
authors.affiliation: Max von Pettenkofer-Institut für Hygiene und Medizinische Mikrobiologie, Ludwig-Maximilians-Universität, München, Germany.|German Center for Infection Research (DZIF), Munich, Germany.
authors.forename: Rainer
authors.initials: R
authors.lastname: Haas
mesh_terms: D001426:Bacterial Proteins / Q000032:analysis*; D020013:Calcium Signaling / Q000187:drug effects*; D002478:Cells, Cultured; D004721:Endoplasmic Reticulum / Q000737:chemistry; D006056:Golgi Apparatus / Q000737:chemistry; D016480:Helicobacter pylori / Q000276:immunology / Q000502:physiology*; D054884:Host-Pathogen Interactions; D006801:Humans; D057131:Immune Evasion; D008565:Membrane Proteins / Q000378:metabolism; D009363:Neoplasm Proteins / Q000378:metabolism; D021381:Protein Transport; D000071737:Stromal Interaction Molecule 1; D013601:T-Lymphocytes / Q000382:microbiology*; D014617:Vacuoles / Q000737:chemistry*
chemical_list: D001426:Bacterial Proteins; D008565:Membrane Proteins; D009363:Neoplasm Proteins; C103530:STIM1 protein, human; D000071737:Stromal Interaction Molecule 1; C074689:VacA protein, Helicobacter pylori