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Journal of Cancer Research and Clinical Oncology

Erschienen in:

01.06.2010 | Original Paper

Nucleolin as cell surface receptor for tumor necrosis factor-α inducing protein: a carcinogenic factor of Helicobacter pylori

verfasst von: Tatsuro Watanabe, Hideaki Tsuge, Takahito Imagawa, Daisuke Kise, Kazuya Hirano, Masatoshi Beppu, Atsushi Takahashi, Kensei Yamaguchi, Hirota Fujiki, Masami Suganuma

Erschienen in: Journal of Cancer Research and Clinical Oncology | Ausgabe 6/2010

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Abstract

Purpose

Tumor necrosis factor-α inducing protein (Tipα) is a unique carcinogenic factor released from Helicobacter pylori (H. pylori). Tipα specifically binds to cells and is incorporated into cytosol and nucleus, where it strongly induces expression of TNF-α and chemokine genes mediated through NF-κB activation, resulting in tumor development. To elucidate mechanism of action of Tipα, we studied a binding protein of Tipα in gastric epithelial cells.

Methods

Tipα binding protein was found in cell lysates of mouse gastric cancer cell line MGT-40 by FLAG-pull down assay and identified to be cell surface nucleolin by flow cytometry using anti-nucleolin antibody. Incorporation of Tipα into the cells was determined by Western blotting and expression of TNF-α gene was quantified by RT-PCR.

Results

Nucleolin was co-precipitated with Tipα-FLAG, but not with del-Tipα-FLAG (an inactive mutant). After treatment with Tipα-FLAG, incorporated Tipα was co-immunoprecipitated with endogenous nucleolin using anti-nucleolin antibody. The direct binding of Tipα to recombinant His-tagged nucleolin fragment (284–710) was also confirmed. Although nucleolin is an abundant non-ribosomal protein of the nucleolus, we found that nucleolin is present on the cell surface of MGT-40 cells. Pretreatment with anti-nucleolin antibody enhanced Tipα-incorporation into the cells through nucleolin internalization. In addition, pretreatment with tunicamycin, an inhibitor of N-glycosylation, decreased the amounts of cell surface nucleolin and inhibited both internalization of Tipα and expression of TNF-α gene.

Conclusions

All the results indicate that nucleolin acts as a receptor for Tipα and shuttles Tipα from cell surface to cytosol and nuclei. These findings provide a new mechanistic insight into gastric cancer development with Tipα.

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Titel: Nucleolin as cell surface receptor for tumor necrosis factor-α inducing protein: a carcinogenic factor of Helicobacter pylori
verfasst von: Tatsuro Watanabe
Hideaki Tsuge
Takahito Imagawa
Daisuke Kise
Kazuya Hirano
Masatoshi Beppu
Atsushi Takahashi
Kensei Yamaguchi
Hirota Fujiki
Masami Suganuma
Publikationsdatum: 01.06.2010
Verlag: Springer-Verlag
Erschienen in: Journal of Cancer Research and Clinical Oncology / Ausgabe 6/2010
Print ISSN: 0171-5216
Elektronische ISSN: 1432-1335
DOI: https://doi.org/10.1007/s00432-009-0733-y

Springer Medizin

Nucleolin as cell surface receptor for tumor necrosis factor-α inducing protein: a carcinogenic factor of Helicobacter pylori