CK2 phosphorylation stimulates SIRT1 activity.

(A) Deacetylase activity of recombinant WT or 4A SIRT1 that was phosphorylated with CK2 in the presence or absence of TBCA (20 µM). Deacetylase activity of SIRT1 was visualized by immunoblotting with acetylated p53 (K382) antibody. The total level of p53 is visualized by Ponceau staining the total level of SIRT1 is visualized by immunoblotting with SIRT1 antibody. (B) The deacetylase activity of the endogenous SIRT1 in the presence of siRNA for either a scrambled sequence or for both catalytic subunits of CK2. P53 K382 acetylation was visualized 2 and 7 hr after IR (20Gy) in U2OS cells. Total levels of p53, SIRT1, CK2α, CK2α' and actin are shown below. (C) H1299 cells co-expressing V5-tagged WT or 4A SIRT1 and p53 in the presence or absence of TBCA were harvested 6 hr after IR (20 Gy). Acetylation of p53 was visualized by immunoblotting with acetylated p53 (K382) antibody. (D) V5-tagged WT or 4A SIRT1 was transiently co-expressed with p53 and either HA-tagged WT or KD CK2α' in H1299 cells. Cells were harvested 6 hr after IR and deacetylation of p53 was visualized by immunoblotting with acetylated p53 (K382) antibody. (E) V5-tagged WT or 4A SIRT1 was transiently co-expressed with FLAG-RelA/p65 (a component of NF-κB) and either HA-tagged WT or KD CK2α' in H1299 cells. FLAG-RelA/p65 was immunoprecipitated with FLAG antibody and immunoblotted with either FLAG antibody to visualize the total level of FLAG-RelA/p65 or Ac-Lys antibody to visualize the acetylation level.