FIG. 8.

α-Tubulin is fully acetylated in primary HDAC6 knockout MEFs. (A) Immunodepletion of acetyl-tubulin was performed with extracts from wild-type (WT) and HDAC6 knockout (KO) primary MEFs. Input (IN) (lanes 1 and 4), supernatant (S) (lanes 2 and 5), and pellet (P) (lanes 3 and 6) fractions were analyzed by SDS-polyacrylamide gel electrophoresis followed by immunoblotting (Western blotting [WB]) with an antibody against acetylated α-tubulin or against α-tubulin; in each case, an amount equivalent to 4% of the initial sample was loaded. IP, immunoprecipitation. (B) Percentage of tubulin acetylation in MEFs and mouse organs. Quantification of tubulin acetylation level in wild-type and HDAC6 knockout primary MEFs, testis, spleen, and brain extracts. Immunoblot quantification of serial extract dilutions was done as described in Materials and Methods; the level of acetylation in the knockout sample was set at 100%. Data of one representative experiment are presented (n = 2 or 3).