title: Molecular Basis of the Ligand Binding Specificity of αvβ8 Integrin.
abstract: αvβ8 is an integrin that recognizes an Arg-Gly-Asp (RGD) motif and interacts with fibronectin, vitronectin, and latent TGF-β1. We comprehensively determined the binding activity of the αvβ8 integrin toward 25 secreted proteins having an RGD motif. The αvβ8 integrin strongly bound to latent TGF-β1 but showed marginal activity for other RGD-containing proteins, including fibronectin and vitronectin. Site-directed mutagenesis of latent TGF-β1 demonstrated that the high affinity binding of αvβ8 integrin to latent TGF-β1 was defined by Leu-218 immediately following the RGD motif within the latency-associated peptide of TGF-β1. Consistent with the critical role of Leu-218 in latent TGF-β1 recognition by αvβ8 integrin, a 9-mer synthetic peptide containing an RGDL sequence strongly inhibited interactions of latent TGF-β1 with αvβ8 integrin, whereas a 9-mer peptide with an RGDA sequence was ∼60-fold less inhibitory. Because αvβ3 integrin did not exhibit strong binding to latent TGF-β1 or distinguish between RGDL- and RGDA-containing peptides, we explored the mechanism by which the integrin β8 subunit defines the high affinity binding of latent TGF-β1 by αvβ8 integrin. Production of a series of swap mutants of integrin β8 and β3 subunits indicated that the high affinity binding of αvβ8 integrin with latent TGF-β1 was ensured by interactions between the Leu-218 residue and the β8 I-like domain, with the former serving as an auxiliary recognition residue defining the restricted ligand specificity of αvβ8 integrin toward latent TGF-β1. In support of this conclusion, high affinity binding toward the αvβ8 integrin was conferred on fibronectin by substitution of its RGDS motif with an RGDL sequence.
authors.affiliation: From the Institute for Protein Research, Osaka University, 3-2 Yamadaoka, Suita, Osaka 565-0871, Japan.
authors.forename: Akio
authors.initials: A
authors.lastname: Ozawa
authors.affiliation: From the Institute for Protein Research, Osaka University, 3-2 Yamadaoka, Suita, Osaka 565-0871, Japan.
authors.forename: Yuya
authors.initials: Y
authors.lastname: Sato
authors.affiliation: From the Institute for Protein Research, Osaka University, 3-2 Yamadaoka, Suita, Osaka 565-0871, Japan.
authors.forename: Tsukasa
authors.initials: T
authors.lastname: Imabayashi
authors.affiliation: From the Institute for Protein Research, Osaka University, 3-2 Yamadaoka, Suita, Osaka 565-0871, Japan.
authors.forename: Toshihiko
authors.initials: T
authors.lastname: Uemura
authors.affiliation: From the Institute for Protein Research, Osaka University, 3-2 Yamadaoka, Suita, Osaka 565-0871, Japan.
authors.forename: Junichi
authors.initials: J
authors.lastname: Takagi
authors.affiliation: From the Institute for Protein Research, Osaka University, 3-2 Yamadaoka, Suita, Osaka 565-0871, Japan sekiguch@protein.osaka-u.ac.jp.
authors.forename: Kiyotoshi
authors.initials: K
authors.lastname: Sekiguchi
mesh_terms: D015153:Blotting, Western; D002448:Cell Adhesion; D002478:Cells, Cultured; D006801:Humans; D016023:Integrins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008024:Ligands; D016297:Mutagenesis, Site-Directed; D009154:Mutation / Q000235:genetics; D009842:Oligopeptides / Q000737:chemistry / Q000378:metabolism*; D010446:Peptide Fragments; D011487:Protein Conformation; D013379:Substrate Specificity; D053773:Transforming Growth Factor beta1 / Q000737:chemistry / Q000378:metabolism*
chemical_list: D016023:Integrins; D008024:Ligands; D009842:Oligopeptides; D010446:Peptide Fragments; D053773:Transforming Growth Factor beta1; C403414:integrin alphavbeta8; C047981:arginyl-glycyl-aspartic acid
keywords: RGD motif; cell adhesion; extracellular matrix; fibronectin; integrin; transforming growth factor beta (TGF-β)