SIR-2.4 interacts with DAF-16 and promotes DAF-16 deacetylation and function independently of catalytic activity.

<p>(A) <i>sir-2.4</i> deletion promotes DAF-16 hyperacetylation. DAF-16 acetylation was assessed in control or <i>sir-2.4</i> KO worms by acetyl-lysine immunoprecipitation followed by GFP immunoblot. (B) SIR-2.4 and DAF-16 interact. Plasmids encoding FLAG-tagged SIR-2.4 and HA-tagged DAF-16 were transfected into 293T cells as indicated (GFP, negative control). Immunoprecipitation and immunoblotting were performed as shown. (C) Rescue of DAF-16 nuclear localization with a catalysis-defective <i>sir-2.4</i> mutant. Stable transgenic strains of <i>sir-2.4(n5137)</i> were generated expressing either wild-type SIR-2.4 or the <i>sir-2.4</i> N124A mutant. Worms were scored for GFP accumulation within the head hypodermic nuclei as day 1 adult (n = 50 or greater) after 20 min of heat-shock at 35°C. <i>P</i>-values were calculated by Pearson's chi-square test.