FIG 2.

Testing the acetylated-K376 G3BP1-specific affinity-purified rabbit polyclonal antibody. (A) 293T and G3BP1-null 293T cells were left untreated or were treated with the deacetylase inhibitors TSA, sodium butyrate, and nicotinamide overnight before lysate preparation. Immunoblotting of the extracts was performed with the indicated antibodies. (B) G3BP1-null 293T cells were transfected with FLAG-tagged G3BP1, G3BP2A, and G3BP2B expression constructs or FLAG vector control. A day later, the cells were treated with the deacetylase inhibitors TSA, sodium butyrate, and nicotinamide overnight before lysate preparation, followed by FLAG immunoprecipitation and immunoblotting with the indicated antibodies. Representative results from at least three independent experiments are shown. (C) The anti-acetylated K376 G3BP1 antibody (Ac-G3BP1) was unable to detect the acetylated G3BP1 band in DACi-treated 293T lysate when the antibody was blocked by preincubation with the immunizing peptide at a 1-μg/ml concentration for 1 h at room temperature.