title: Interaction of cartilage oligomeric matrix protein/thrombospondin 5 with aggrecan.
abstract: Cartilage oligomeric matrix protein/thrombospondin 5 (COMP/TSP5) is a major component of the extracellular matrix (ECM) of the musculoskeletal system. Its importance is underscored by its association with several growth disorders. In this report, we investigated its interaction with aggrecan, a major component of cartilage ECM. We also tested a COMP/TSP5 mutant, designated MUT3 that accounts for 30% of human pseudoachondroplasia cases, to determine if the mutation affects function. Using a solid-phase binding assay, we have shown that COMP/TSP5 can bind aggrecan. This binding was decreased with MUT3, or when COMP/TSP5 was treated with EDTA, indicating the presence of a conformation-dependent aggrecan binding site. Soluble glycosaminoglycans (GAGs) partially inhibited binding, suggesting that the interaction was mediated in part through aggrecan GAG side chains. Using affinity co-electrophoresis, we showed that COMP/TSP5, in its calcium-replete conformation, bound to heparin, chondroitin sulfates, and heparan sulfate; this binding was reduced with EDTA treatment of COMP/TSP5. MUT3 showed weaker binding than calcium-repleted COMP/TSP5. Using recombinant COMP/TSP5 fragments, we found that the "signature domain" could bind to aggrecan, suggesting that this domain can mediate the interaction of COMP/TSP5 and aggrecan. In summary, our data indicate that COMP/TSP5 is an aggrecan-binding protein, and this interaction is regulated by the calcium-sensitive conformation of COMP/TSP5; interaction of COMP with aggrecan can be mediated through the GAG side chains on aggrecan and the "signature domain" of COMP/TSP5. Our results suggest that COMP/TSP5 may function to support matrix interactions in cartilage ECM.
authors.affiliation: Cartilage Biology and Orthopaedics Branch, NIAMS, National Institutes of Health, Bethesda, Maryland 20892, USA. chenfl@mail.nih.gov
authors.forename: Faye Hui
authors.initials: FH
authors.lastname: Chen
authors.forename: Mary E
authors.initials: ME
authors.lastname: Herndon
authors.forename: Nichlesh
authors.initials: N
authors.lastname: Patel
authors.forename: Jacqueline T
authors.initials: JT
authors.lastname: Hecht
authors.forename: Rocky S
authors.initials: RS
authors.lastname: Tuan
authors.forename: Jack
authors.initials: J
authors.lastname: Lawler
mesh_terms: D053674:Aggrecans / Q000378:metabolism*; D000818:Animals; D001665:Binding Sites; D002356:Cartilage / Q000378:metabolism; D064236:Cartilage Oligomeric Matrix Protein; D002809:Chondroitin Sulfates / Q000737:chemistry; D004330:Drosophila; D004492:Edetic Acid / Q000737:chemistry; D016326:Extracellular Matrix Proteins / Q000378:metabolism*; D006023:Glycoproteins / Q000378:metabolism*; D006025:Glycosaminoglycans / Q000378:metabolism; D006493:Heparin / Q000378:metabolism; D006801:Humans; D007700:Kinetics; D064235:Matrilin Proteins; D008954:Models, Biological; D011485:Protein Binding; D017434:Protein Structure, Tertiary
chemical_list: D053674:Aggrecans; D064236:Cartilage Oligomeric Matrix Protein; D016326:Extracellular Matrix Proteins; D006023:Glycoproteins; D006025:Glycosaminoglycans; D064235:Matrilin Proteins; C577553:TSP5 protein, human; D006493:Heparin; D002809:Chondroitin Sulfates; D004492:Edetic Acid