Altered synthesis of cartilage-specific proteoglycans by mutant human cartilage oligomeric matrix protein.
BACKGROUND
The mechanism by which mutant cartilage oligomeric matrix protein (COMP) induces a pseudoachondroplasia phenotype remains unknown, and the reason why a mutation of a minor protein of the growth plate cartilage causes total disruption of endochondral bone formation has not yet been determined. The current study was performed to investigate the effects of mutated COMP on the synthesis of the cartilage-specific major matrix proteins of Swarm rat chondrosarcoma chondrocytes.


METHODS
The Swarm rat chondrosarcoma chondrocytes transfected with a chimeric construct, which consisted of a mutant gene of human COMP and an amino acid FLAG tag sequence, were cultured in agarose gel. Formation of extracellular proteoglycan and type-II collagen by the cells was evaluated by immunohistochemical staining and measuring the (35)S-sulfate incorporation.


RESULTS
No difference was observed for the detection of type-II collagen among the cell lines expressing mutant COMP and the control cell lines. Histochemical staining of sulfated proteoglycans with safranin-O showed that lesser amounts of proteoglycans were incorporated into the extracellular matrix of the chondrocytes transfected with the mutant gene. (35)S-sulfate incorporation into the cell/matrix fractions demonstrated markedly lower radiolabel incorporation, as compared to that of the control cells.


CONCLUSIONS
Mutation of COMP has an important impact on the processing of proteoglycans, rather than type-II collagen, in the three-dimensional culture of Swarm rat chondrosarcoma chondrocytes.
Kwak
Yoon Hae
YH
Department of Orthopaedic Surgery, Hallym University Sacred Heart Hospital, Anyang, Korea.
Roh
Jae Young
JY
Lee
Ki Seok
KS
Park
Hui Wan
HW
Kim
Hyun Woo
HW
D053674:Aggrecans / Q000032:analysis / Q000096:biosynthesis*; D000818:Animals; D064236:Cartilage Oligomeric Matrix Protein; D002478:Cells, Cultured; D019902:Chondrocytes / Q000378:metabolism*; D002813:Chondrosarcoma / Q000378:metabolism; D024043:Collagen Type II / Q000096:biosynthesis*; D005109:Extracellular Matrix / Q000378:metabolism*; D016326:Extracellular Matrix Proteins / Q000235:genetics*; D006023:Glycoproteins / Q000235:genetics*; D006801:Humans; D064235:Matrilin Proteins; D009154:Mutation; D051381:Rats; D014162:Transfection
D053674:Aggrecans; D064236:Cartilage Oligomeric Matrix Protein; D024043:Collagen Type II; D016326:Extracellular Matrix Proteins; D006023:Glycoproteins; D064235:Matrilin Proteins; C577553:TSP5 protein, human