Isolation, Cloning, and Sequence Analysis of the Integrin Subunit α10, a β1-associated Collagen Binding Integrin Expressed on Chondrocytes

Abstract

We have found that chondrocytes express a novel collagen type II-binding integrin, a new member of the ␤1-integrin family. The integrin ␣ subunit, which has a M r of 160 kDa reduced, was isolated from bovine chondrocytes by collagen type II affinity purification. The human homologue was obtained by screening a human chondrocyte library with a bovine cDNA probe. Cloning and cDNA sequence analysis of the human integrin ␣ subunit designated ␣10 show that it shares the general structure of other integrin ␣ subunits. The predicted amino acid sequence consists of a 1167-amino acid mature protein, including a signal peptide (22 amino acids), a long extracellular domain (1098 amino acids), a transmembrane domain (25 amino acids), and a short cytoplasmic domain (22 amino acids). The extracellular part contains a 7-fold repeated sequence, an I-domain (199 amino acids) and three putative divalent cation-binding sites. The deduced amino acid sequence of ␣10 is 35% identical to the integrin subunit ␣2 and 37% identical to the integrin subunit ␣1. Northern blot analysis shows a single mRNA of 5.4 kilobases in chondrocytes. A peptide antibody against the predicted sequence of the cytoplasmic domain of ␣10 immunoprecipitated two proteins with masses of 125 and 160 kDa from chondrocyte lysates under reducing conditions. The peptide antibody specifically stained chondrocytes in tissue sections of human articular cartilage, showing that ␣10␤1 is expressed in cartilage tissue.The integrins are a large family of transmembrane glycoproteins that mediate cell-cell and cell-matrix interactions (1-5). All known members of this superfamily are noncovalently associated heterodimers composed of an ␣ and a ␤ subunit. At present, 8 ␤-(␤1-␤8) (See Ref. 6 and references therein) and 16 ␣ subunits (␣1-␣9, ␣v, ␣M, ␣L, ␣X, ␣IIb, ␣E, and ␣D) have been characterized (6 -21), and these subunits associate to generate more than 20 different integrins. The ␤1 subunit has been shown to associate with 10 different ␣ subunits, ␣1-␣9 and ␣v and to mediate interactions with extracellular matrix proteins such as collagens, laminins, and fibronectin. The major collagen binding integrins are ␣1␤1 and ␣2␤1 (22-25). The integrins ␣3␤1 and ␣9␤1 have also been reported to interact with collagen (26, 27), although this interaction is not well understood (28). The extracellular N-terminal regions of the ␣ and ␤ integrin subunits are important in the binding of ligands (29,30). The N-terminal region of the ␣ subunits is composed of a 7-fold repeated sequence (12, 31) containing FG and GAP consensus sequences. The repeats are predicted to fold into a ␤-propeller domain (32), with the last three or four repeats containing putative divalent cation binding sites. The ␣-integrin subunits ␣1, ␣2, ␣D, ␣E, ␣L, ␣M, and ␣X contain an ϳ200 amino acid inserted domain, the I-domain (A-domain), that shows similarity to sequences in von Willebrand factor, cartilage matrix protein, and complement factors C2 and B (33, 34). The I-domain is localized between the second and third FG-GA...

Cathepsins B, K, and L Are Regulated by a Defined Collagen Type II Peptide via Activation of Classical Protein Kinase C and p38 MAP Kinase in Articular Chondrocytes

“…Alternatively to the annexin link, ␣10␤1 integrin function may also be affected by the treatment with collagen peptides. This integrin has been shown to bind to triple-helical collagen type II (55), and it may also interfere with the same signaling pathway. Putative strategies to further clarify the role of the peptide feedback loop in chondrocytes to control ECM turnover will have to consider the various options.…”

Effects of I Domain Deletion on the Function of the β2 Integrin Lymphocyte Function-associated Antigen-1

“…A subset of nine integrins incorporates an additional, autonomously folding domain of ϳ200 amino acids, which is inserted between ␤-sheets 2 and 3 of the putative ␤-propeller and is termed the I (inserted) domain. The I domain is present in LFA-1 and the other ␤2 integrins Mac-1, p150,95, and ␣d␤2, as well as in ␣1␤1, ␣2␤1, ␣10␤1, ␣11␤1, and ␣E␤7 (Camper et al, 1998;Dickeson and Santoro, 1998;Velling et al, 1999). The crystal structures of the I domains of LFA-1, Mac-1, and ␣2␤1 have been solved and show a dinucleotidebinding fold (reviewed by Loftus and Liddington, 1997;Humphries and Newham, 1998).…”

Collagen XXIII, Novel Ligand for Integrin α2β1 in the Epidermis

“…This ␤ 1 subunit-containing subclass of integrins is characterized by the presence of an inserted domain (I domain) with homology to von Willebrand factor A domains in their ␣-subunit, which harbors the ligand-binding site (8,9). The integrins ␣ 10 ␤ 1 and ␣ 11 ␤ 1 have a restricted expression pattern on differentiated chondrocytes and developing muscle cells (10,11), whereas ␣ 1 ␤ 1 and ␣ 2 ␤ 1 show a broader distribution, with integrin ␣ 1 ␤ 1 being predominantly expressed by cells of mesenchymal origin and integrin ␣ 2 ␤ 1 by epithelial cells and platelets. In vitro studies suggested an implication of integrin ␣ 2 ␤ 1 in cell attachment and migration (12,13), generation of mechanical forces and contraction of collagen matrices (14), induction of collagenase activity and matrix remodeling (15,16), as well as angiogenesis (17) and epithelial branching morphogenesis (18).…”