Tyrosine-phosphorylation of the 12th armadillo-repeat of beta-catenin is associated with cadherin dysfunction in human cancer.
Tyrosine phosphorylation of beta-catenin, an intracytoplasmic cadherin-binding protein, causes disruption of the cadherin-mediated cell adhesion system in cancer cells. We identified that the c-erbB-2 protein activated by TGFalpha was capable of binding to the 12th armadillo repeat (arm12) of beta-catenin with increased phosphorylation-state level. We produced a monoclonal antibody, 4G7, which recognizes a phosphorylated-tyrosine residue (Tyr-654) located at arm12 of beta-catenin. Tyrosine phosphorylation of the arm12 was detected after stimulation with TGFalpha in TMK-1. Immunoprecipitation analyses using 4G7, anti-beta-catenin, alpha-catenin, the c-erbB-2 gene product and phosphotyrosine antibodies indicated that tyrosine phosphorylation of the arm12 was closely correlated with dissociation between E-cadherin/beta-catenin and alpha-catenin or the c-erbB-2 gene product, but not with dissociation between beta-catenin and E-cadherin. Immunocytochemical staining of TMK-1 cells using the 4G7 antibody revealed that tyrosine phosphorylation of the arm12 was localized at cell-cell contact sites of cancer cells transiently and only after TGFalpha treatment. Immunohistochemical localization of the 4G7 antibody was observed in cancer cells at the invasive front where they detached from cancer nests. These findings indicated that the tyrosine phosphorylation of arm12 is a key marker of cadherin dysfunction and the 4G7 antibody may be useful in screening for a beta-catenin phosphorylation ligand or tyrosine kinases.
Shomori
Kohei
K
Pathology Division, National Cancer Center Research Institute, 1-1 Tsukiji 5-chome, Chuo-ku, Tokyo, Japan.
Ochiai
Atsushi
A
Akimoto
Shingo
S
Ino
Yoshinori
Y
Shudo
Koyo
K
Ito
Hisao
H
Hirohashi
Setsuo
S
D000911:Antibodies, Monoclonal; D015153:Blotting, Western; D015820:Cadherins / Q000378:metabolism*; D002448:Cell Adhesion / Q000502:physiology; D002478:Cells, Cultured; D004797:Enzyme-Linked Immunosorbent Assay; D005455:Fluorescent Antibody Technique; D006801:Humans; D007150:Immunohistochemistry; D047468:Immunoprecipitation; D009361:Neoplasm Invasiveness / Q000235:genetics* / Q000473:pathology; D010766:Phosphorylation; D016133:Polymerase Chain Reaction; D018719:Erb-b2 Receptor Tyrosine Kinases / Q000235:genetics / Q000378:metabolism*; D013274:Stomach Neoplasms / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D016211:Transforming Growth Factor alpha / Q000378:metabolism; D014443:Tyrosine / Q000378:metabolism*; D051176:beta Catenin / Q000235:genetics* / Q000276:immunology / Q000378:metabolism
D000911:Antibodies, Monoclonal; D015820:Cadherins; D018719:Erb-b2 Receptor Tyrosine Kinases; D016211:Transforming Growth Factor alpha; D014443:Tyrosine; D051176:beta Catenin