Reduced HDAC6 activity in ATP13A2-null cells. (a–f) Increased α-tubulin acetylation with ATP13A2 deficiency. Lysates of ATP13A2-null (KO) HEK293 cells (a) and mouse liver tissues (c), ATP13A2 RNAi Drosophila brain tissues (e) and their corresponding controls (WT) were immunodetected for acetyl-α-tubulin (Ac-tub), HDAC6, and α-tubulin (α-tub). Respective quantitation of Ac-tub is shown (b, d, and f). Two independent ATP13A2-null cell lines (a; KO1 and -2), liver tissues from three ATP13A2-null mice (c; KO 1, 2, and 3), and three independent Drosophila ATP13A2 RNAi lines (e, HMS02719, GD14627, and HMS05242) were analyzed. For Drosophila, RNAi expression was driven by elav-gal4 (e; Elav>). An elav-gal4 line served as control (e; Elav/+). *, P < 0.05; **, P < 0.01, n = 3. (g and h) Reduced HDAC6 activity in ATP13A2-null cells. ATP13A2-null HEK293 (KO) and control cells (WT) were treated with HDAC6 inhibitor tubacin for 16 h followed by tubacin removal. Cells were collected at indicated time points (post wash). Cell lysates were immunodetected for acetyl-α-tubulin (g; Ac-tub) and α-tubulin (α-tub). Relative levels of Ac-tub (Ac-tub/α-tub) are shown (h). *, P < 0.05, n = 3.