Isolation of a novel integrin receptor mediating Arg-Gly-Asp-directed cell adhesion to fibronectin and type I collagen from human neuroblastoma cells. Association of a novel beta 1-related subunit with alpha v.
We report the isolation from two human neuroblastoma cell lines of an Arg-Gly-Asp-dependent integrin complex capable of binding to vitronectin, fibronectin, and type I collagen. The two neuroblastoma cell lines, SK-N-SH and IMR-32, exhibit specific attachment to fibronectin and type I collagen. SK-N-SH cells exhibit a much stronger attachment to vitronectin than the IMR-32 cells, which attach poorly to this substrate. Affinity chromatography of octylglucoside extracts of 125I surface-labeled cells on GRGDSPK-Sepharose columns resulted in the specific binding and elution with GRGDSP of three radiolabeled polypeptides with relative molecular masses of 135, 115, and 90 kD when analyzed by SDS-PAGE under nonreducing conditions. In the SK-N-SH cells the 135- and 90-kD polypeptides were more abundant whereas in the IMR-32 cells the 135- and 115-kD polypeptides were more highly expressed. Liposomes prepared from fractions containing all three polypeptides bound to vitronectin, fibronectin, and type I collagen, whereas liposomes prepared from the 135- and 115-kD polypeptides bound only to fibronectin and type I collagen. Polyclonal antibodies against the alpha/beta complexes of both the vitronectin receptor and the fibronectin receptor immunoprecipitated all three polypeptides. A monoclonal antibody against beta 1 immunoprecipitated only the 135- and the 115-kD polypeptides, whereas a monoclonal antibody against beta 3 subunit immunoprecipitated the 135- and 90-kD polypeptides. Although, the 115-kD polypeptide could be recognized by an anti-beta 1 antibody, a comparison of peptide maps generated by V8 protease digestion of the 115-kD polypeptide and beta 1 subunit immunoprecipitated from GRGDSPK-Sepharose flow-through material indicated that these two polypeptides are distinct. Depletion of the 90-kD polypeptide with an anti-beta 3 monoclonal antibody did not effect the ability of the 115- and 135-kD polypeptides to bind to GRGDSPK-Sepharose. These data indicate that the SK-N-SH and IMR-32 neuroblastoma cells express a novel "beta 1-like" integrin subunit that can associate with alpha v and can bind to RGD. We propose to name this beta 1-like subunit beta n. The data reported here thus demonstrate that in these two cell lines alpha v associates with two beta subunits, beta n and beta 3, forming two heterodimers. The alpha v beta n complex mediates binding to fibronectin and type I collagen, whereas the alpha v beta 3 complex mediates binding to vitronectin.
Department of Advanced Therapeutics, Cancer Control Agency of British Columbia, Vancouver, Canada.
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D002448:Cell Adhesion / Q000187:drug effects / Q000502:physiology; D003094:Collagen / Q000378:metabolism*; D003110:Colonic Neoplasms / Q000378:metabolism / Q000473:pathology / Q000648:ultrastructure; D005109:Extracellular Matrix / Q000378:metabolism / Q000648:ultrastructure; D005353:Fibronectins / Q000378:metabolism*; D006801:Humans; D016023:Integrins / Q000302:isolation & purification* / Q000378:metabolism / Q000502:physiology; D009447:Neuroblastoma / Q000378:metabolism / Q000473:pathology* / Q000648:ultrastructure; D010975:Platelet Aggregation Inhibitors / Q000378:metabolism / Q000494:pharmacology*; D011971:Receptors, Immunologic / Q000378:metabolism / Q000502:physiology*; D018000:Receptors, Peptide; D019030:Receptors, Vitronectin; D014407:Tumor Cells, Cultured
D005353:Fibronectins; D016023:Integrins; D010975:Platelet Aggregation Inhibitors; D011971:Receptors, Immunologic; D018000:Receptors, Peptide; D019030:Receptors, Vitronectin; C051382:arginyl-glycyl-aspartic acid directed cell adhesion receptor; D003094:Collagen