title: Cartilage matrix protein forms a type II collagen-independent filamentous network: analysis in primary cell cultures with a retrovirus expression system.
abstract: Cartilage matrix protein (CMP) is expressed specifically in mature cartilage and consists of two von Willebrand factor A domains (CMP-A1 and CMP-A2) that are separated by an epidermal growth factor-like domain, and a coiled-coil tail domain at the carboxyl terminal end. We have shown previously that CMP interacts with type II collagen-containing fibrils in cartilage. In this study, we describe a type II collagen-independent CMP filament and we analyze the structural requirement for the formation of this type of filament. Recombinant wild-type CMP and two mutant forms were expressed in chick primary cell cultures using a retrovirus expression system. In chondrocytes, the wild-type virally encoded CMP is able to form disulfide bonded trimers and to assemble into filaments. Filaments also form with CMP whose Cys455 and Cys457 in the tail domain were mutagenized to prevent interchain disulfide bond formation. Therefore, intermolecular disulfide bonds are not necessary for the assembly of CMP into filaments. Both the wild-type and the double cysteine mutant also form filaments in fibroblasts, indicating that chondrocyte-specific factors are not required for filament formation. A truncated form of CMP that consists only of the CMP-A2 domain and the tail domain can form trimers but fails to form filaments, indicating that the deleted CMP-A1 domain and/or the epidermal growth factor domain are necessary for filament assembly but not for trimer formation. Furthermore, the expression of the virally encoded truncated CMP in chondrocyte culture disrupts endogenous CMP filament formation. Together these data suggest a role for CMP in cartilage matrix assembly by forming filamentous networks that require participation and coordination of individual domains of CMP.
authors.affiliation: Cutaneous Biology Research Center, Massachusetts General Hospital, Charlestown 02129, USA.
authors.forename: Q
authors.initials: Q
authors.lastname: Chen
authors.forename: D M
authors.initials: DM
authors.lastname: Johnson
authors.forename: D R
authors.initials: DR
authors.lastname: Haudenschild
authors.forename: M M
authors.initials: MM
authors.lastname: Tondravi
authors.forename: P F
authors.initials: PF
authors.lastname: Goetinck
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002356:Cartilage / Q000166:cytology / Q000502:physiology* / Q000648:ultrastructure; D002478:Cells, Cultured; D002642:Chick Embryo; D003094:Collagen / Q000502:physiology; D003545:Cysteine; D017931:DNA Primers; D016326:Extracellular Matrix Proteins; D005347:Fibroblasts / Q000166:cytology / Q000502:physiology / Q000648:ultrastructure; D005822:Genetic Vectors; D006023:Glycoproteins / Q000096:biosynthesis / Q000502:physiology*; D064235:Matrilin Proteins; D008954:Models, Biological; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D017354:Point Mutation; D016133:Polymerase Chain Reaction; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000378:metabolism; D012190:Retroviridae; D012694:Serine; D014162:Transfection
chemical_list: D017931:DNA Primers; D016326:Extracellular Matrix Proteins; D006023:Glycoproteins; D064235:Matrilin Proteins; D011994:Recombinant Proteins; D012694:Serine; D003094:Collagen; D003545:Cysteine