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Papers In Press, published online ahead of print June 13, 2001
J. Biol. Chem, 10.1074/jbc.M011442200
Submitted on December 19, 2000
Revised on June 13, 2001
Accepted on June 13, 2001
University of Nantes, Lab. Physiopathologie de la résorption osseuse, Nantes cedex 1 44035
Corresponding Author: francoise.redini{at}sante.univ-nantes.fr
Decorin, a small leucin-rich proteoglycan may play an important role in the attempt of cartilage repair initiated by chondrocytes in early stages of osteoarthritis, through its ability to bind collagen fibrils and growth factors such as TGF-b. We previously demonstrated that TGF-b decreased decorin mRNA steady state levels in articular chondrocytes (Demoor, M., Rédini, F., Boittin, M., and Pujol, J.-P. (1998) Biochim Biophys Acta 1398, 179-191). Here, we investigated the effect of TGF-b on decorin gene expression in both primary cultures of articular chondrocytes and chondrocytes dedifferentiated by serial passages. Transient transfection of cells with plasmid constructs of decorin promoter linked to the luciferase reporter gene revealed transcriptional repression by TGF-b, in fully differentiated as well as dedifferentiated chondrocytes. Experiments with 5' deleted constructs allowed characterization of a TGF-b responsive element in the shortest construct (-155/+269 bp). DNase I footprinting analysis delineated a negative TGF-b responsive region between -140 and -111 bp in the decorin proximal promoter. Gel-retardation assays demonstrated that TGF-b modulates decorin gene expression through transcription factors the nature and mode of action of which are depending on the differentiation state of the chondrocytes : two DNA-protein complexes were formed in the region -144/-127 bp with nuclear extracts from primary chondrocytes, whereas a higher mobility complex was observed in the -127/-111 bp region for dedifferentiated cells. Antibodies against vitamin D and retinoic acid receptors used in supershift experiments showed that these nuclear receptors are involved in the regulation of decorin gene expression in articular chondrocytes.
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