FIG. 8.

Inhibition of Sox9-mediated activation of the Col2a1 enhancer by CRYBP1. RCS and NIH 3T3 cells were transiently transfected with 2 μg of the reporter construct pKN159Bx6luc containing six tandem copies of the Col2a1 enhancer sequences along with 2 μg of the C-ZF expression vector, Sox9 expression vector, or control vector (pCA1F). C-ZF suppressed the enhancer activity of pKN159Bx6luc to 32% of that of the control in RCS cells. pKN159Bx6 was not active in NIH 3T3 cells; however, expression of Sox9 resulted in significant activation of the Col2a1 enhancer in the nonchondrocytic cells. Cotransfection of increasing amounts of the C-ZF expression vector inhibits Sox9-mediated activation of the Col2a1 enhancer in a dose-dependent manner. A Renilla luciferase expression vector, pRL-SV40, was used as an internal control for transfection efficiency. The relative luciferase activities are average values ± the standard errors for three independent transfected cultures from two experiments.