FIGURE 3.

FoxO1 is required for nitric oxide-induced GADD45α expression. A, INS 832/13 cells were treated with DEANO (1 mm) for 1 h followed by washing and 5-h incubation in the absence of DEANO. The comet assay was used to examine DNA damage. B and C, INS 832/13 cells were transduced with Δ256-FoxO1 or WT-FoxO1 adenoviral constructs. Twenty-four hours after adenoviral transduction the cells were treated with DEANO (1 mm) for 1 h, 3 h, or 1 h followed by wash and 5-h recovery incubation. Total RNA was isolated, and real time PCR for GADD45α (B) or hsp70 (C) was performed and normalized to actin levels. D, alternatively, after adenoviral transduction, FoxO1 expression was examined by Western blot analysis. E and F, INS 832/13 cells transduced with Δ256-FoxO1 or WT-FoxO1 adenoviral constructs were treated with DEANO (1 mm) for 1 h or treated for DEANO for 1 h followed by washing and 5-h recovery incubation. The comet assay was used to assess DNA damage, and representative comet tails are shown (E) or quantified as the mean tail moment (F). Results are the average ± S.E. (error bars) of three independent experiments (A, B, C, and F; or representative of three independent experiments (D and E). Statistical significance is indicated (*, p < 0.05 compared with untreated control, †, p < 0.05 compared with WT-FoxO1 transduced cells).