TPA-induced cohort migration of well-differentiated human rectal adenocarcinoma cells: cells move in a RGD-dependent manner on fibronectin produced by cells, and phosphorylation of E-cadherin/catenin complex is induced independently of cell-extracellular matrix interactions.
We have already presented a two-dimensional cell motility assay using a highly metastatic variant (L-10) of human rectal adenocarcinoma cell line RCM-1 as a motility model of tumour cells of epithelial origin. In this model, L-10 cells showed locomotion as a coherent sheet when stimulated with 12-O-tetradecanoylphorbol-13-acetate (TPA), and we called this type of movement "cohort migration". Electron and immunoelectron microscopic study of the migrating cell sheets demonstrated localized release from cell-cell adhesion only at the lower portion of the cells with loss of E-cadherin immunoreactivity, and this change was associated with increased tyrosine phosphorylation of the E-cadherin-catenin complex, including beta-catenin. Cell-extracellular matrix (ECM) interactions involved in this TPA-induced cohort migration and their effect on tyrosine phosphorylation of the E-cadherin-catenin complex have now been investigated. L-10 cell cohort migration was almost completely inhibited by addition of Arg-Gly-Asp (RGD) peptide into the medium, and thus RGD dependent. Cohort migration was stimulated on type I and IV collagens, fibronectin (FN)- and laminin-coated substratum, but was inhibited by RGD only on FN-coated surface. By using immunofluorescent techniques, FN was demonstrated preferentially around migrating cells, and a protein synthesis inhibitor, cycloheximide, inhibited the migration by about 75%. FN produced by L-10 cells were found to be mostly EDA+ FN when analysed by RT-PCR. Moreover, anti-FN antibody, but not anti-vitronectin antibody, inhibited the TPA-induced cohort migration almost completely. Thus, it was likely that L-10 cells produced FN themselves and moved on the FN substrate in an RGD-dependent manner. However, stimulation of migration by type I collagen coating and inhibition by RGD treatment did not affect the tyrosine phosphorylation of the E-cadherin-catenin complex induced by TPA, indicating that cell-cell interactions were adjusted to suit cell migration, irrespective of the condition of cell-ECM adhesion, during TPA-induced cohort migration.
Department of Pathology, Miyazaki Medical College, Kiyotake, Japan.
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K
Nabeshima
T
T
Inoue
Y
Y
Shimao
H
H
Kataoka
M
M
Koono
D000230:Adenocarcinoma / Q000378:metabolism / Q000473:pathology*; D000911:Antibodies, Monoclonal / Q000494:pharmacology; D015820:Cadherins / Q000378:metabolism*; D002448:Cell Adhesion; D002450:Cell Communication; D002465:Cell Movement / Q000187:drug effects* / Q000502:physiology; D003598:Cytoskeletal Proteins / Q000378:metabolism*; D017931:DNA Primers / Q000737:chemistry; D005109:Extracellular Matrix / Q000378:metabolism; D005353:Fibronectins / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D016253:Microscopy, Immunoelectron; D009842:Oligopeptides / Q000494:pharmacology*; D010766:Phosphorylation; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000378:metabolism; D012004:Rectal Neoplasms / Q000378:metabolism / Q000473:pathology*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology*; D015534:Trans-Activators; D014407:Tumor Cells, Cultured; D051176:beta Catenin
D000911:Antibodies, Monoclonal; C495270:CTNNB1 protein, human; D015820:Cadherins; D003598:Cytoskeletal Proteins; D017931:DNA Primers; D005353:Fibronectins; D009842:Oligopeptides; D012333:RNA, Messenger; D015534:Trans-Activators; D051176:beta Catenin; C047981:arginyl-glycyl-aspartic acid; D013755:Tetradecanoylphorbol Acetate