Figure 6
From: Notch signaling controls chondrocyte hypertrophy via indirect regulation of Sox9
Notch signaling inhibits Sox9 gene expression via secondary effectors. (a) Diagram for the localization of a RBPjk consensus binding site (yellow box is wild-type sequence and blue box is mutant sequence) and N-box consensus binding site (red box) in the Sox9 promoter and luciferase constructs. Core consensus sequences are listed in diagram. (b) ATDC5 cells were co-transfected with Flag or NICD1 over-expression vectors and either a wild-type 6.8 kb Sox9-Luciferase complex (−6.8 kb WT) or 6.8 kb construct with a mutated RBPjk binding site (−6.8 kb MT). Luciferase levels were measured 24 hours after transfection. (c) ATDC5 cells were co-transfected with Flag or NICD1 over-expression vectors and either a 1 kb, 0.5 kb, or 0.32 kb Sox9-Luciferase deletion constructs. Luciferase levels were measured 24 hours after transfection. (d) Quantitative RT-PCR assessing Hes1 (d1, d2), Hey1 (d3, d4), and Sox9 (d5, d6) gene expression in ATDC5 cells at 0-, 2-, 4-, and 8-hour post-culture on IgG versus JAG1 coated plates in the absence (d1, d3, d5) or presence (d2, d4, d6) of cycloheximide.
