FIG 6.

SIRT1 represses RTA transactivation in reporter assays. (A to F) SIRT1 represses RTA activation of its own promoter (A and B) and the vIL-6 promoter (C and D) but not LANA latent promoter LTc (E and F), as shown in reporter assays (A, C, and E) together with Western blots of the expressed RTA and SIRT1 proteins (B, D, and F). 293T cells were cotransfected with the luciferase reporter plasmid pRp-luc, pvIL-6-luc, or pLTc-luc together with RTA or without RTA in the presence of increased doses of a SIRT1 expression plasmid. A β-galactosidase expression construct was used to calibrate the transfection efficiency. The luciferase activity of the reporter without RTA and SIRT1 was given a value of 1. The protein extracts were also examined by Western blotting to detect the expression of the RTA and SIRT1 proteins. (G and H) NAM relieves SIRT1's inhibitory effect on RTA transactivation of the vIL-6 promoter. A reporter assay was carried out as described in the legend to panel A. NAM (10 mM) was added at 5 h posttransfection. The protein extracts were also examined by Western blotting to detect the expression of the RTA and SIRT1 proteins.