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...

Chondroadherin (the 36-kD protein) is a leucine-rich, cartilage matrix protein known to mediate adhesion of isolated chondrocytes. In the present study we investigated cell surface proteins involved in the interaction of cells with chondroadherin in cell adhesion and by affinity purification. Adhesion of bovine articular chondrocytes to chondroadherin-coated dishes was dependent on Mg2+ or Mn2+ but not Ca2+. Adhesion was partially inhibited by an antibody recognizing β1 integrin subunit. Chondroadherin-binding proteins from chondrocyte lysates were affinity purified on chondroadherin-Sepharose. The β1 integrin antibody immunoprecipitated two proteins with molecular mass ∼110 and 140 kD (nonreduced) from the EDTA-eluted material. These results indicate that a β1 integrin on chondrocytes interacts with chondroadherin. To identify the α integrin subunit(s) involved in interaction of cells with the protein, we affinity purified chondroadherin-binding membrane proteins from human fibroblasts. Immunoprecipitation of the EDTA-eluted material from the affinity column identified α2β1 as a chondroadherin-binding integrin. These results are in agreement with cell adhesion experiments where antibodies against the integrin subunit α2 partially inhibited adhesion of human fibroblast and human chondrocytes to chondroadherin. Since α2β1 also is a receptor for collagen type II, we tested the ability of different antibodies against the α2 subunit to inhibit adhesion of T47D cells to collagen type II and chondroadherin. The results suggested that adhesion to collagen type II and chondroadherin involves similar or nearby sites on the α2β1 integrin. Although α2β1 is a receptor for both collagen type II and chondroadherin, only adhesion of cells to collagen type II was found to mediate spreading.

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