SIRT1 interacts with RORγt. (A) Flag-tagged WT or H363Y mutant SIRT1 was immunoprecipitated from transfected 293T cells and probed as indicated. (B) RORγt was immunoprecipitated from thymocytes and Th17 cells, and probed with antibody against SIRT1. (C) Immunoprecipitation using lysates of 293T cells co-transfected with constructs encoding SIRT1 and various deletion mutants of RORγt. Relative binding was calculated by normalizing the ratio of immunoprecipitated RORγt/SIRT1 to the ratio of input RORγt/SIRT1. (D) Acetylation of Flag-tagged RORγt immunoprecipitated from 293T cells transfected with various acetyltransferases and Flag-RORγt. (E and F) Acetylation of Flag-RORγt co-transfected with p300 and WT or H363Y mutant SIRT1, in the absence (E) or in the presence (F) of nicotinamide and Ex-527. Equal amounts of Flag-RORγt were loaded (D–F). Representative data are shown from four (A), three (B and E), and two (D) independent experiments, and combined data are shown from three (C and F) independent experiments with error bars representing ±SEM.
SIRT1 interacts with RORγt. (A) Flag-tagged WT or H363Y mutant SIRT1 was immunoprecipitated from transfected 293T cells and probed as indicated. (B) RORγt was immunoprecipitated from thymocytes and Th17 cells, and probed with antibody against SIRT1. (C) Immunoprecipitation using lysates of 293T cells co-transfected with constructs encoding SIRT1 and various deletion mutants of RORγt. Relative binding was calculated by normalizing the ratio of immunoprecipitated RORγt/SIRT1 to the ratio of input RORγt/SIRT1. (D) Acetylation of Flag-tagged RORγt immunoprecipitated from 293T cells transfected with various acetyltransferases and Flag-RORγt. (E and F) Acetylation of Flag-RORγt co-transfected with p300 and WT or H363Y mutant SIRT1, in the absence (E) or in the presence (F) of nicotinamide and Ex-527. Equal amounts of Flag-RORγt were loaded (D–F). Representative data are shown from four (A), three (B and E), and two (D) independent experiments, and combined data are shown from three (C and F) independent experiments with error bars representing ±SEM.
