Figure S2.

CHN-1/CHIP Overexpression Affects Lifespan, Related to Figure 2

(A) Level of CHN-1::FLAG overexpression strain was analyzed by immunoblotting of indicated worm lysates (day 1 and 10 old animals).

(B) Overexpression of chn-1 does not enhance lifespan of wild-type worms (n = 4 independent experiments).

(C) Overexpression of dCHIP was driven by the use of the midgut specific TIGS-2 driver line and dCHIP levels were verified in gut extracts. Equal loading was verified by ponceau S staining (ponceau).

(D) Flies of the genotype TIGS-2/ UAS-dCHIP were either placed on ethanol or RU486 (1 μg/ml or 5 μg/ml) containing food to induce dCHIP expression. dCHIP expression leads to toxicity in a dose-dependent manner and results in reduced lifespan.

(E) Depletion of CHIP in HEK293 cells does not affect the expression level of PDK1, AKT, mTOR, or Hsp90.

(F) Specificity of anti-DAF-2 antibodies was evaluated using daf-2(e1370) worms expressing thermolabile DAF-2 protein. Worms were grown at 25°C until day 1 of adulthood, lysed, and analyzed by western blotting using DAF-2 specific antibodies.