title: The CD9, CD81, and CD151 EC2 domains bind to the classical RGD-binding site of integrin αvβ3.
abstract: Tetraspanins play important roles in normal (e.g. cell adhesion, motility, activation, and proliferation) and pathological conditions (e.g. metastasis and viral infection). Tetraspanins interact with integrins and regulate integrin functions, but the specifics of tetraspanin-integrin interactions are unclear. Using co-immunoprecipitation with integrins as a sole method to detect interaction between integrins and full-length tetraspanins, it has been proposed that the variable region (helices D and E) of the extracellular-2 (EC2) domain of tetraspanins laterally associates with a non-ligand-binding site of integrins. We describe that, using adhesion assays, the EC2 domain of CD81, CD9, and CD151 bound to integrin αvβ3, and this binding was suppressed by cRGDfV, a specific inhibitor of αvβ3, and antibody 7E3, which is mapped to the ligand-binding site of β3. We also present evidence that the specificity loop of β3 directly bound to the EC2 domains. This suggests that the EC2 domains specifically bind to the classical ligand-binding site of αvβ3. αvβ3 was a more effective receptor for the EC2 domains than the previously known tetraspanin receptors α3β1, α4β1, and α6β1. Docking simulation predicted that the helices A and B of CD81 EC2 bind to the RGD-binding site of αvβ3. Substituting Lys residues at positions 116 and 144/148 of CD81 EC2 in the predicted integrin-binding interface reduced the binding of CD81 EC2 to αvβ3, consistent with the docking model. These findings suggest that, in contrast with previous models, the ligand-binding site of integrin αvβ3, a new tetraspanin receptor, binds to the constant region (helices A and B) of the EC2 domain.
authors.affiliation: Department of Dermatology, Biochemistry and Molecular Medicine, University of California Davis School of Medicine, Sacramento, CA 95817, U.S.A.|Ph.D. Program for Translational Medicine, College of Medical Sciences and Technology, Taipei Medical University, Taipei, Taiwan.
authors.forename: Jessica
authors.initials: J
authors.lastname: Yu
authors.affiliation: Ph.D. Program for Translational Medicine, College of Medical Sciences and Technology, Taipei Medical University, Taipei, Taiwan.
authors.forename: Chia-Ying
authors.initials: CY
authors.lastname: Lee
authors.affiliation: Ph.D. Program for Translational Medicine, College of Medical Sciences and Technology, Taipei Medical University, Taipei, Taiwan.|Ph.D. Program for Cancer Biology and Drug Discovery, College of Medical Science and Technology, Taipei Medical University, Taipei, Taiwan.|Core Facility, Taipei Medical University, Taipei, Taiwan.
authors.forename: Chun Austin
authors.initials: CA
authors.lastname: Changou
authors.affiliation: Department of Dermatology, Biochemistry and Molecular Medicine, University of California Davis School of Medicine, Sacramento, CA 95817, U.S.A.
authors.forename: Dora M
authors.initials: DM
authors.lastname: Cedano-Prieto
authors.affiliation: Department of Dermatology, Biochemistry and Molecular Medicine, University of California Davis School of Medicine, Sacramento, CA 95817, U.S.A.|Ph.D. Program for Translational Medicine, College of Medical Sciences and Technology, Taipei Medical University, Taipei, Taiwan.
authors.forename: Yoko K
authors.initials: YK
authors.lastname: Takada
authors.affiliation: Department of Dermatology, Biochemistry and Molecular Medicine, University of California Davis School of Medicine, Sacramento, CA 95817, U.S.A. ytakada@ucdavis.edu ytakada@tmu.edu.tw.|Ph.D. Program for Translational Medicine, College of Medical Sciences and Technology, Taipei Medical University, Taipei, Taiwan.
authors.forename: Yoshikazu
authors.initials: Y
authors.lastname: Takada
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000737:chemistry; D001665:Binding Sites; D016466:CHO Cells; D003001:Cloning, Molecular; D003412:Cricetulus; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D015870:Gene Expression; D039302:Integrin alphaVbeta3 / Q000737:chemistry* / Q000235:genetics / Q000276:immunology; D007700:Kinetics; D062105:Molecular Docking Simulation; D011485:Protein Binding; D000072756:Protein Conformation, alpha-Helical; D000072757:Protein Conformation, beta-Strand; D054730:Protein Interaction Domains and Motifs; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D060185:Tetraspanin 24 / Q000737:chemistry* / Q000235:genetics / Q000276:immunology; D060225:Tetraspanin 28 / Q000737:chemistry* / Q000235:genetics / Q000276:immunology; D060245:Tetraspanin 29 / Q000737:chemistry* / Q000235:genetics / Q000276:immunology
chemical_list: D000911:Antibodies, Monoclonal; C554715:CD151 protein, human; C554798:CD81 protein, human; C554869:CD9 protein, human; D039302:Integrin alphaVbeta3; D011994:Recombinant Proteins; D060185:Tetraspanin 24; D060225:Tetraspanin 28; D060245:Tetraspanin 29
keywords: CD151; CD81; CD9; RGD-binding site; integrins; tetraspanins