FIGURE 1.

Identification of the acetylation sites of TIP60. A, schematic of various truncated TIP60 proteins fused with GST and putative acetylated sites of TIP60 protein. aa, amino acids; FL, full-length. B, various GST-TIP60 fusion proteins were subjected to an in vitro acetylation assay. Top panel, representative radioautography data. Bottom panel, gel code blue staining. C, schematic representation of various point mutant constructs of TIP60. D and E, HEK293 cells were transfected with wild-type or mutant FLAG-tagged TIP60 constructs for 24 h and then incubated with 1 μm TSA and 5 mm nicotinamide for an additional 6 h. The acetylation levels of TIP60 (Ac-TIP60) and total TIP60 protein were detected after anti-FLAG immunoprecipitation. The polyclonal antibody against acetylated lysine (Cell Signaling Technology, catalog no. 9441) was employed in these experiments.