title: Effect of ligand conformation on melanoma cell alpha3beta1 integrin-mediated signal transduction events: implications for a collagen structural modulation mechanism of tumor cell invasion.
abstract: The importance of three-dimensional interactions between receptors with their respective ligands has been extensively explored during the binding process, but considerably less so for postbinding events such as induction of signaling pathways. Tumor cell receptor association with basement membrane proteins is believed to facilitate the metastatic process. Melanoma and ovarian carcinoma cells have been shown to utilize the alpha3beta1 integrin to bind to models of the alpha1(IV)531-543 sequence from basement membrane (type IV) collagen [Miles, A. J., et al. (1994) J. Biol. Chem. 269, 30939-30945; Miles, A. J., et al. (1995) J. Biol. Chem. 270, 29047-29050]. In the present study, the effects of ligand three-dimensional structure on possible signal transduction pathways induced by alpha3beta1 integrin binding have been evaluated. Human melanoma cell binding to type IV collagen resulted in Tyr phosphorylation of p125(FAK), consistent with prior studies correlating beta1 integrin subunit binding to collagen and p125(FAK) Tyr phosphorylation. Cross-linking of an anti-alpha3 integrin subunit monoclonal antibody also induced p125(FAK) Tyr phosphorylation. Incubation of melanoma cells with single-stranded or triple-helical peptide models of alpha1(IV)531-543 induced Tyr phosphorylation of intracellular proteins. Immunoprecipitation analysis identified one of these proteins as pp125(FAK). Induction of p125(FAK) Tyr phosphorylation was enhanced and the time of induction was shortened when the ligand was used in triple-helical conformation. Subsequent clustering of either the single-stranded or the triple-helical ligand also increased the level of p125(FAK) phosphorylation compared to unclustered ligand. The clustered triple-helical peptide ligand induced more rapid paxillin Tyr phosphorylation than the single-stranded ligand. In addition, the induction of activated proteases was found to be more rapid due to ligand triple helicity. Overall, these studies have shown that (i) a model of an isolated sequence from type IV collagen, alpha1(IV)531-543, can induce alpha3beta1 integrin-mediated signal transduction in melanoma cells and (ii) ligand conformation (secondary, tertiary, and/or quaternary structure) can directly influence several alpha3beta1 integrin-mediated signal transduction events. The effects of ligand conformation suggest that a "collagen structural modulation" mechanism may exist for tumor cell invasion, whereby triple-helical collagen promotes cell binding and induction of signal transduction, subsequently leading to collagen dissolution by proteases, decreased signal transduction, and enhanced tumor cell motility.
authors.affiliation: Department of Chemistry and Biochemistry, Florida Atlantic University, Boca Raton, Florida 33431, USA.
authors.forename: J L
authors.initials: JL
authors.lastname: Lauer
authors.forename: C M
authors.initials: CM
authors.lastname: Gendron
authors.forename: G B
authors.initials: GB
authors.lastname: Fields
mesh_terms: D002448:Cell Adhesion; D015815:Cell Adhesion Molecules / Q000378:metabolism; D003094:Collagen / Q000737:chemistry / Q000378:metabolism*; D051417:Focal Adhesion Kinase 1; D051416:Focal Adhesion Protein-Tyrosine Kinases; D006801:Humans; D039201:Integrin alpha3beta1; D016023:Integrins / Q000378:metabolism*; D008024:Ligands; D008545:Melanoma / Q000378:metabolism / Q000473:pathology*; D008954:Models, Biological; D009361:Neoplasm Invasiveness; D011233:Precipitin Tests; D011485:Protein Binding; D011487:Protein Conformation; D011505:Protein-Tyrosine Kinases / Q000378:metabolism; D015398:Signal Transduction; D014407:Tumor Cells, Cultured
chemical_list: D015815:Cell Adhesion Molecules; D039201:Integrin alpha3beta1; D016023:Integrins; D008024:Ligands; D003094:Collagen; D011505:Protein-Tyrosine Kinases; D051417:Focal Adhesion Kinase 1; D051416:Focal Adhesion Protein-Tyrosine Kinases; C496050:PTK2 protein, human