Figure 3

Effect of HDAC6 on A3G expression and substrate deacetylation. a Left, western blot of dose–response effects of EGFP-wt-HDAC6 and EGFP-HDAC6-ΔBUZ (detected by anti-HDAC6 or anti-EGFP abs) on A3G-3xHA expression. Right, quantitation of western blots regarding EGFP-HDAC6- and EGFP-HDAC6-ΔBUZ-mediated effect on A3G expression. All experiments were performed in HEK 293T cells. When indicated, endogenous HDAC6 expression level is shown. All western blots are representative of three independent experiments. b Western blot of EGFP-wt-HDAC6-mediated Hsp90 deacetylation in HEK 293T cells in a dose–response assay. Acetylated (Ac.) Hsp90 was detected from endogenous Hsp90 pulled-down from cell lysates and with an anti-Hsp90 Ab, then blotted with a specific anti-acetylated Lys Ab. c Western blot of EGFP-wt-HDAC6-mediated α-tubulin deacetylation (compare lanes 1 and 3) and inhibition of HIV-1 Env (rs-gp120_IIIB)-induced acetylation of α-tubulin (compare lanes 2 and 4), in a CD4 + T CEM.NKR-CCR5 cell line. In b, c, EGFP-wt-HDAC6 and acetylated (Ac.) α-tubulin were detected by anti-EGFP and anti-acetylated α-tubulin Abs, respectively. From a to c, α-tubulin is the control for total protein. When indicated, endogenous HDAC6 expression level is shown. All western blots are representative of three independent experiments.