Profiling of the tetraspanin web of human colon cancer cells.
Tetraspanins are integral membrane proteins involved in a variety of physiological and pathological processes. In cancer, clinical and experimental studies have reported a link between tetraspanin expression levels and metastasis. Tetraspanins play a role as organizers of multimolecular complexes in the plasma membrane. Indeed each tetraspanin associates specifically with one or a few other membrane proteins forming primary complexes. Thus, tetraspanin-tetraspanin associations lead to a molecular network of interactions, the "tetraspanin web." We performed a proteomic characterization of the tetraspanin web using a model of human colon cancer consisting of three cell lines derived from the primary tumor and two metastases (hepatic and peritoneal) from the same patient. The tetraspanin complexes were isolated after immunoaffinity purification using monoclonal antibodies directed against the tetraspanin CD9, and the associated proteins were separated by SDS-PAGE and identified by mass spectrometry using LC-MS/MS. This allowed the identification of 32 proteins including adhesion molecules (integrins, proteins with Ig domains, CD44, and epithelial cell adhesion molecule) (EpCAM), membrane proteases (ADAM10, TADG-15, and CD26/dipeptidyl peptidase IV), and signaling proteins (heterotrimeric G proteins). Importantly some components were differentially detected in the tetraspanin web of the three cell lines: the laminin receptor Lutheran/B-cell adhesion molecule (Lu/B-CAM) was expressed only on the primary tumor cells, whereas CD26/dipeptidyl peptidase IV and tetraspanin Co-029 were observed only on metastatic cells. Concerning Co-029, immunohistofluorescence showed a high expression of Co-029 on epithelial cells in normal colon and a lower expression in tumors, whereas heterogeneity in terms of expression level was observed on metastasis. Finally we demonstrated that epithelial cell adhesion molecule and CD9 form a new primary complex in the tetraspanin web.
Le Naour
François
F
INSERM U602, Institut André Lwoff, Université Paris XI, Hôpital Paul Brousse, 94807 Villejuif Cedex, France. flenaour@vjf.inserm.fr
André
Magali
M
Greco
Céline
C
Billard
Martine
M
Sordat
Bernard
B
Emile
Jean-François
JF
Lanza
François
F
Boucheix
Claude
C
Rubinstein
Eric
E
D015703:Antigens, CD / Q000378:metabolism*; D000951:Antigens, Neoplasm / Q000378:metabolism; D015815:Cell Adhesion Molecules / Q000378:metabolism; D002853:Chromatography, Liquid; D003110:Colonic Neoplasms / Q000737:chemistry / Q000378:metabolism* / Q000473:pathology*; D000071858:Epithelial Cell Adhesion Molecule; D006801:Humans; D007413:Intestinal Mucosa / Q000166:cytology / Q000473:pathology; D013058:Mass Spectrometry; D008562:Membrane Glycoproteins / Q000378:metabolism*; D008565:Membrane Proteins / Q000737:chemistry; D009362:Neoplasm Metastasis; D011485:Protein Binding; D021381:Protein Transport; D040901:Proteomics; D060245:Tetraspanin 29; D059470:Tetraspanins; D014407:Tumor Cells, Cultured
D015703:Antigens, CD; D000951:Antigens, Neoplasm; C554869:CD9 protein, human; D015815:Cell Adhesion Molecules; D000071858:Epithelial Cell Adhesion Molecule; D008562:Membrane Glycoproteins; D008565:Membrane Proteins; C065609:TSPAN8 protein, human; D060245:Tetraspanin 29; D059470:Tetraspanins