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Helicobacter pylori CagA Containing ITAM‐Like Sequences Localized to Lipid Rafts Negatively Regulates VacA‐Induced Signaling In Vivo
Author(s) -
Asahi Momoyo,
Tanaka Yukie,
Izumi Toshimasa,
Ito Yoshiyuki,
Naiki Hironobu,
Kersulyte Dangeruta,
Tsujikawa Kazutake,
Saito Masakazu,
Sada Kiyonao,
Yanagi Shigeru,
Fujikawa Akihiro,
Noda Masaharu,
Itokawa Yoshinori
Publication year - 2003
Publication title -
helicobacter
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.206
H-Index - 79
eISSN - 1523-5378
pISSN - 1083-4389
DOI - 10.1046/j.1523-5378.2003.00118.x
Subject(s) - caga , tyrosine phosphorylation , phosphorylation , tyrosine , receptor tyrosine kinase , microbiology and biotechnology , biology , protein tyrosine phosphatase , sh2 domain , helicobacter pylori , tyrosine kinase , biochemistry , lipid raft , chemistry , signal transduction , genetics , virulence , gene
Background. Helicobacter pylori CagA is injected into the host cell and tyrosine‐phosphorylated. We examined tyrosine‐phosphorylation sites of CagA, as well as the function of CagA proteins in vivo and in vitro. Methods. After proteolytic digestion of CagA with lysyl endopeptidase, CagA tyrosine‐phosphorylation sites were determined using quadropolar time‐of‐flight (Q‐TOF) mass spectrometry analysis. Specific anti‐pY CagA polyclonal and anti‐CagA monoclonal antibodies were used to examine gastric mucosal biopsy specimens from H. pylori infected patients. Results. Mass spectrometry identified five crucial tyrosine‐phosphorylation sites of CagA at Tyr893, Tyr912, Tyr965, Tyr999, and Tyr1033 within the five repeated EPIYA sequences of H. pylori (NCTC11637)‐infected AGS cells. CagA protein also had an immuno‐receptor tyrosine‐based activation motif (ITAM)‐like amino acid sequences in the 3′ region of the cagA , E PI Y AT I x 27 EI Y AT I , which closely resembled the ITAM. CagA proteins: (i) were localized to the 1% TritonX‐100 resistant membrane fraction (lipid rafts); (ii) formed a cluster of phosphorylated CagA protein complexes; (iii) associated with tyrosine‐phosphorylated GIT1/Cat1 (G protein‐coupled receptor kinase‐interactor 1/Cool‐associated tyrosine‐phosphorylated 1), substrate molecules of receptor type protein‐tyrosine phosphatase (RPTPζ/β), which is the receptor of VacA; and (iv) were involved in a delay and negative regulation of VacA‐induced signal. Furthermore, immunohistochemical staining of gastric mucosal biopsy specimens provided strong evidence that tyrosine‐phosphorylated CagA is found together with CagA at the luminal surface of gastric foveola in vivo. Conclusion. These findings suggest an important role for CagA containing ITAM‐like sequences in the pathogenesis of H. pylori‐ related disease.