FIGURE 6.
The selective repression of CSIG, PCNA, and CCNA2 by Rb is TAAC element-dependent. A, U2OS cells cotransfected with vectors alone (Ctrl) or p16 and shRNA against Rb (shRb) with CSIG, PCNA, or CCNA2 promoter luciferase reporter constructs or their TAAC element mutants or deletions. CSIG TAAC mut, TAAC to TGAC mutant of the CSIG promoter; PCNA TAAC mut, TAAC to TGAC mutant of the PCNA promoter; CCNA2 TAAC del, the fragment of the CCNA2 promoter with a TAAC element deletion. The values represent the means and standard errors for three independent experiments. B, U2OS cells cotransfected with vectors alone (Ctrl) or with p16 and E2F1 overexpression vectors with CSIG promoter luciferase reporter constructs or the TAAC element mutant were analyzed for luciferase expression. The values represent the means and standard errors for three independent experiments. C, U2OS cells cotransfected with vectors alone (Ctrl) or p16 and shRNA against Rb (shRb) with ARF and p27KIP1 promoter luciferase reporter constructs or their TAAC insertion mutants. Insertion of the TAAC element into p27 and ARF promoters was constructed by introducing a TAAC element into the −244-bp position of the p27 promoter and the −478-bp position of the ARF promoter. The values represent the means and standard errors for three independent experiments. D, U2OS cells cotransfected with vectors alone (Ctrl) or p16 and shRNA against Rb (shRb) together with wild-type CSIG promoter luciferase reporter constructs or their E2F binding site mutant. The values represent the means and standard errors for three independent experiments. Ctrl, control.