The mechanism of cleavage of EGFR ligands induced by inflammatory cytokines in gastric cancer cells.

Abstract

BACKGROUND & AIMS:

The epidermal growth factor (EGF) receptor (EGFR) can be transactivated by many factors including G-protein-coupled receptor agonists and cytokines. Although this EGFR transactivation reportedly requires a disintegrin and metalloproteinase (ADAM) that sheds the ectodomain of EGFR ligands, the detailed mechanisms are still unknown. This study evaluated the mechanism of interleukin (IL)-8- and IL-1beta-dependent shedding of the EGFR ligand in KATO III cells.

METHODS:

We established transfectants stably expressing alkaline phosphatase-tagged heparin-binding EGF-like growth factor (HB-EGF), transforming growth factor alpha, or amphiregulin precursors, and depleted ADAM proteins, using short interfering RNA against ADAM10, 12, or 17. We assessed shedding of EGFR ligands by measuring AP activities in the conditioned media after IL-1beta or IL-8 stimulation. EGFR activation was examined by immunoprecipitation and Western blotting using antiphosphotyrosine antibody. KB-R7785 and anti-IL-8 neutralizing antibody were used to inhibit activities of ADAMs and IL-8 action, respectively.

RESULTS:

IL-8 dose dependently released the EGFR ligands and transiently phosphorylated EGFR, with a peak at 15 minutes. KB-R7785 completely blocked IL-8-induced shedding and EGFR transactivation. Depletion of ADAM10 also dramatically reduced IL-8-induced shedding and EGFR transactivation, but depletion of ADAM12 and 17 did not. IL-1beta dose dependently enhanced shedding of HB-EGF, which was not blocked by KB-R7785 in the early phase. In the late phase, however, the EGFR transactivation was blocked by KB-R7785 and abrogated by anti-IL-8 neutralizing antibody.

CONCLUSIONS:

IL-8 induces shedding of EGFR ligands because of an ADAM10-dependent pathway in gastric cancer cells, whereas IL-1beta acts principally by an ADAM-independent pathway. IL-1beta-dependent prolonged EGFR transactivation involves multiple pathways, including an IL-8-dependent pathway.

• Authors: Satoshi Tanida, Takashi Joh, Keisuke Itoh, Hiromi Kataoka, Makoto Sasaki, Hirotaka Ohara, Takahiro Nakazawa, Tomoyuki Nomura, Yumi Kinugasa, Hiroshi Ohmoto, Hiroshi Ishiguro, Kohichiro Yoshino, Shigeki Higashiyama, Makoto Itoh
• Journal: Gastroenterology (Gastroenterology) Vol. 127 Issue 2 Pg. 559-69 (Aug 2004) ISSN: 0016-5085 [Print] United States
• PMID: 15300588 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)

Chemical References

• Antibodies
• Interleukin-1
• Interleukin-8
• Ligands
• Membrane Proteins
• RNA, Small Interfering
• Tyrosine
• ErbB Receptors
• Amyloid Precursor Protein Secretases
• ADAM Proteins
• Metalloendopeptidases
• ADAM10 Protein
• ADAM10 protein, human

Topics

• ADAM Proteins
• ADAM10 Protein
• Amyloid Precursor Protein Secretases
• Antibodies (pharmacology)
• Cell Line, Tumor
• ErbB Receptors (immunology, metabolism)
• Humans
• Interleukin-1 (genetics, metabolism)
• Interleukin-8 (genetics, metabolism)
• Ligands
• Membrane Proteins (chemistry, genetics, metabolism)
• Metalloendopeptidases (chemistry, genetics, metabolism)
• Phosphorylation
• Protein Structure, Tertiary
• RNA, Small Interfering
• Stomach Neoplasms
• Tyrosine (metabolism)