SP3 represses the SP1-mediated transactivation of the human COL2A1 gene inprimary and de-differentiated chondrocytes

Sp1 and Sp3 effects on the transcription of the human alpha1(II) procollage n gene (COL2A1) were investigated in both differentiated and de-differentia ted rabbit articular chondrocytes. Transient transfection with constructs o f deleted COL2A1 promoter sequences driving the luciferase reporter gene re vealed that the region spanning -266 to +121 base pairs showed Sp1-enhancin g effects, whatever the differentiation state. In contrast, Sp3 did not inf luence COL2A1 gene transcription. Concomitant overexpression of the two Sp proteins demonstrated that Sp3 blocked the Sp1 induction of COL2A1 promoter activity. Moreover, inhibition of Sp1/Sp3 binding to their target DNA sequ ence decreased both COL2A1 gene transcription and Sp1-enhancing effects. DN ase I footprinting and gel retardation assays revealed that Sp1 and Sp3 bin d specifically to cis-sequences of the COL2A1 gene promoter whereby they ex ert their transcriptional effects. Sp1 and Sp3 levels were found to be redu ced in de-differentiated chondrocytes, as revealed by DNA-binding and immun ochemical study. Sp1 specifically activated collagen neosynthesis whatever the differentiation state of chondrocytes, suggesting that this factor exer ts a major role in the expression of collagen type II. However, our data in dicate that type II collagen-specific expression in chondrocytes depend on both the Sp1/Sp3 ratio and cooperation of Sp1 with other transcription fact ors, the amounts of which are also modulated by phenotype alteration.