Fig. 5: p27 promotes microtubule acetylation.

A Acetylated (Ac-) α-tubulin immunostaining of p27^+/+ and p27^−/− MEFs in full medium or glucose-deprived for 24 h and 48 h. B Quantification of acetylated tubulin intensity normalized to the mean value in p27^+/+ cells in full medium from experiments described in (A). Number of cells used for quantification: p27^+/+: n = 388 (0 h), n = 474 (24 h), n = 706 (48 h); p27^−/−: n = 492 (0 h), n = 1316 (24 h), n = 689 (48 h). C Ac-α-tubulin and α-tubulin immunoblots in p27^+/+ and p27^−/− MEFs in full medium or glucose deprived for 48 h. D p27 immunoblot in p27^−/− MEFs infected with either empty vector or p27. α-tubulin was used as loading control. E p27^−/− MEFs infected with either empty vector or p27 were glucose-starved for 48 h and immunostained for Ac-α-tubulin and α-tubulin. DNA was stained with Hoechst 33342. F Quantification of acetylated tubulin fluorescence intensity in p27^−/− MEFs re-expressing p27 normalized to control vector infected cells from n = 3 independent experiments as described in (E). At least 155 cells were analyzed per condition per experiment. G Ac-α-tubulin, LC3B and α-tubulin immunostaining of p27^−/− MEFs infected with either empty vector or p27, glucose starved for 48 h. A, E, G Scale bars are 50 µm. B, F Graphs show means ± SEM. Statistical significance was determined using 2-way ANOVA (B) or unpaired two-tailed Student’s t test with Welch’s correction (F).