title: Analysis of integrin receptors for laminin and type IV collagen on metastatic B16 melanoma cells.
abstract: As tumor cells invade surrounding tissue, they adhere to various extracellular matrix components. Previously we reported that B16-BL6 melanoma cell adhesion to both basement membrane and purified protein substrates was blocked by antibody to beta 1-integrin adhesion receptors (R. H. Kramer et al., Cancer Res., 49: 393-402, 1989). In the present study we found, using immunofluorescent staining, that beta 1-integrin complexes were colocalized with vinculin in focal adhesion plaques on laminin, type IV collagen, and fibronectin substrates. To identify potential adhesion receptors on B16 cells, the cells were surface-labeled with 125I, solubilized with detergent, and chromatographed on laminin-, type IV collagen-, and fibronectin-Sepharose columns. On laminin-Sepharose, an integrin heterodimer complex was eluted with EDTA that contained a beta 1 chain at Mr 120,000 and an alpha subunit at Mr 140,000 (nonreduced). This complex was specific for laminin and failed to bind to collagen- or fibronectin-Sepharose columns. Immunoprecipitation with specific monoclonal antibody identified this complex as alpha 6 beta 1 (VLA-6). Furthermore, monoclonal antibody to the alpha 6 beta 1 complex effectively blocked the attachment of B16-BL6 cells to laminin but did not affect adhesion to fibronectin or type IV collagen. We recovered a different integrin complex from type IV collagen-Sepharose columns that was composed of a beta 1 chain and an alpha chain of Mr 180,000 (nonreduced). This same complex also exhibited a weak affinity for laminin-affinity chromatography. The laminin-binding complex and the type IV collagen-binding complex were clearly distinct from the fibronectin-binding receptor and were not eluted by arginyl-glycyl-aspartate-containing peptides. The results suggest that the B16 melanoma cells express multiple integrin-related receptors that appear to mediate cell adhesion to basement membrane matrices.
authors: Department of Stomatology, University of California, San Francisco 94143.
authors: D M
authors: DM
authors: Ramos
authors: E D
authors: ED
authors: Berston
authors: R H
authors: RH
authors: Kramer
mesh_terms: D000818:Animals; D002448:Cell Adhesion; D002846:Chromatography, Affinity; D003094:Collagen / Q000378:metabolism*; D005455:Fluorescent Antibody Technique; D016023:Integrins / Q000378:metabolism*; D007526:Isoelectric Point; D007797:Laminin / Q000378:metabolism*; D008546:Melanoma, Experimental / Q000378:metabolism* / Q000473:pathology; D051379:Mice; D008970:Molecular Weight; D009362:Neoplasm Metastasis; D011233:Precipitin Tests; D011956:Receptors, Cell Surface / Q000378:metabolism; D038981:Receptors, Collagen; D017469:Receptors, Fibronectin; D011971:Receptors, Immunologic / Q000378:metabolism; D017474:Receptors, Laminin
chemical_list: D016023:Integrins; D007797:Laminin; D011956:Receptors, Cell Surface; D038981:Receptors, Collagen; D017469:Receptors, Fibronectin; D011971:Receptors, Immunologic; D017474:Receptors, Laminin; D003094:Collagen