FoxO1 deacetylation couples Sirt1 action to POMC expression.

(A) Representative western blots showing the acetylation status of FoxO1 and Stat3 upon fasting and intraperitoneal leptin administration. 5 animals per group were evaluated. (B) Fasting induced FoxO1 deacetylation is Sirt1 dependent. Rats were icv treated with DMSO (fed and fasted groups) or EX-527 as in Figure 1D. ARCs were analyzed for the expression of Ac-FoxO1 and total FoxO1 by western blotting. 5 animals per group were evaluated. (C) Inhibition of Sirt1 activity decreases FoxO1 activity in vivo. (D) Activation of Sirt1 in hypothalamic N43 cell line increases FoxO1 activity. (E) FoxO1 knock-down abrogates the inhibitory effect of Sirt1 on POMC expression. AtT20 cells were transfected with the corresponding siRNAs as described in Figure 3D. 24 hr after transfection the cells were serum starved and treated either with DMSO or SA3 (5 µM) for 6 hours. ACTH in the media was measured by RIA. (F) Hypothalamic knock-down of FoxO1 reverses the affect of Sirt1 inhibition on POMC (*, vs. siControl DMSO group). Values are the mean±SEM. *, vs. respective control groups unless stated otherwise.