Nuclear factor binding to an AP-1 site is associated with the activation of pro-alpha 1(I)-collagen gene in dedifferentiating chondrocytes.
Isolated chondrocytes grown on plastic gradually lose their differentiated phenotype upon subculturing. This dedifferentiation is manifested by an altered production of extracellular-matrix molecules (ECM): e.g., the cartilage specific type II collagen is replaced by types I and III. We have studied the regulation of ECM gene expression in dedifferentiating human and murine fetal chondrocytes. Nuclear extracts from dedifferentiated cells, human fetal fibroblasts and 3T3 cells contained a protein that bound in an electrophoretic mobility shift assay to an AP-1 site in the first intron of the human alpha 1(I) collagen gene. This binding activity was not present in freshly isolated human or murine chondrocytes, which produced type II, but not type I, collagen mRNA in culture. Thus the binding activity was induced simultaneously with alpha 1(I)-collagen-gene expression during dedifferentiation. The specific interaction was sensitive to dephosphorylation of the nuclear extract and to chemical modification of reduced cysteine residues. The AP-1 site we studied had previously been shown to be a positive transcriptional contributor in the first intron to the expression of the alpha 1(I) collagen gene. In transient transfections into dedifferentiating chondrocytes, an alpha 1(I) collagen expression plasmid carrying a mutated AP-1 site in the first intron resulted in three-times-lower reporter gene RNA levels than a plasmid carrying the respective functional AP-1 site. These data suggest that the AP-1 sequence and its respective trans-acting factors may play a role in the transcriptional regulation of the alpha 1(I) collagen gene during dedifferentiation of chondrocytes.
Department of Medical Biochemistry, University of Turku, Finland.
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A
Määttä
V
V
Glumoff
P
P
Paakkonen
D
D
Liska
R P
RP
Penttinen
K
K
Elima
D016475:3T3 Cells / Q000378:metabolism; D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D002356:Cartilage / Q000166:cytology* / Q000378:metabolism*; D002454:Cell Differentiation; D002478:Cells, Cultured; D005333:Fetus; D015870:Gene Expression; D013006:Growth Hormone / Q000235:genetics; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D009154:Mutation; D009687:Nuclear Proteins / Q000378:metabolism*; D010957:Plasmids; D011347:Procollagen / Q000235:genetics*; D016755:Proto-Oncogene Proteins c-jun / Q000378:metabolism*; D012333:RNA, Messenger / Q000378:metabolism; D014162:Transfection
D009687:Nuclear Proteins; D011347:Procollagen; D016755:Proto-Oncogene Proteins c-jun; D012333:RNA, Messenger; D013006:Growth Hormone