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. 2005 Feb 3;24(5):1021–1032. doi: 10.1038/sj.emboj.7600570

Figure 6.

Figure 6

Inhibition of FOXO1 activity by FHL2 requires SIRT1 activity. (A) DU145 cells were transfected with 3 × IRSLuc (0.3 μg) and pCMVβGal (0.3 μg). As indicated, cells were cotransfected with Flag-FOXO1 (0.3 μg) and Myc-FHL2 (0.3 μg). Transfected cells received 5 or 20 mM nicotinamide for 24 h. Luciferase activity was determined and normalized to β-gal activity. Fold activation was calculated by dividing the normalized activity of cells expressing FOXO1 and FHL2 by that of the corresponding vector controls. (B) DU145 cells were transfected with Flag-FOXO1 (3 μg), Myc-FHL2 (3 μg), and 6 μg of SIRT1 or GFP siRNA. Cell extracts were immunoblotted with anti-SIRT1, anti-β-actin, or M2 antibody. (C) DU145 cells were transfected with 3 × IRSLuc (0.3 μg), pRL-null (0.01 μg), Flag-FOXO1 (0.3 μg), and 0.6 μg of the indicated siRNA vectors. Luciferase activity was assayed and normalized as in Figure 1D.