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Exp Cell Res. 1994 Jul;213(1):28-36.

Characterization of type II and type XI collagen synthesis by an immortalized rat chondrocyte cell line (IRC) having a low level of type II collagen mRNA expression.

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  • 1Research Department, Shriners Hospital for Crippled Children, Portland, Oregon 97201.


The biosynthesis of type XI and type II collagens was examined using a stable rat chondrocyte cell line established by W. E. Horton et al. (1988, Exp. Cell Res. 178, 457-468.). These cells (IRC; immortalized rat chondrocytes) were created by transformation with a murine retrovirus carrying the v-myc and v-raf oncogenes. They grow in suspension culture as multicellular aggregates and synthesize typical cartilage proteins, aggrecan and link protein. Type II collagen is absent or synthesized at severely reduced levels, as shown by Northern analysis of mRNA. Thus, this cell type represents a unique model in which to study cartilage matrix protein interactions in the absence of type II collagen. A more detailed look at the proteins secreted into the medium by metabolically labeled IRC cells revealed the presence of collagenase-sensitive bands when analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The bands were identified as the alpha 1, alpha 2, and alpha 3 chains of heterotrimeric type XI collagen by electrophoretic migration after pepsin digestion, by CNBr peptide mapping, and by immunoprecipitation with antibodies to rat alpha 1(XI). mRNA for all three chains was detected by Northern blot analysis. The data indicate that the low level of alpha 1(II) mRNA previously detected in these cells is translated into pro alpha 3(XI) polypeptide chains which are incorporated into molecules of type XI. Under normal culture conditions, homotrimers of type II collagen were not detected. The carboxyl propeptide domain of the fibrillar collagens directs chain selection and molecular assembly of the trimeric molecules. The sequence of the carboxyl propeptide domain from pro alpha 3(XI) of IRC cells was found to be identical to this domain from pro alpha 1(II) of swarm rat chondrosarcoma, supporting previous evidence that pro alpha 3(XI) and pro alpha 1(II) have the same primary structure. When cultured in the presence of 50 mM arginine, IRC cells could be induced to synthesize pro alpha 1(II) chains in excess over pro alpha 1(XI) and pro alpha 2(XI). Only under these conditions were type II collagen molecules detected, suggesting a preferential association of pro alpha 1(II) with the pro alpha 1 and/or pro alpha 2 chains of type XI collagen.

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