Fig. 5

D570Y mutation alters the subcellular distribution and microtubule association of CTTNBP2. a Peculiar distribution of CTTNBP2 D570Y mutant protein in cultured hippocampal neurons at DIV 8. b Average immunoreactivities of WT and D570Y mutant protein along dendrites. c D570Y-expressing neurons possess more dendrites expressing filamentous CTTNBP2. d Increased microtubule association of CTTNBP2 upon D570Y mutation. COS1 cells were separately transfected with WT or D570Y CTTNBP2 and subjected to microtubule spin down assay using microtubule filaments. Immunoblotting using HA tag and α-tubulin antibodies was then performed to analyze the results. Quantification results of four independent experiments are shown in the panel at right. e Increased tubulin acetylation upon expression of D570Y mutant. Cultured hippocampal neurons were cotransfected with GFP and WT or D570Y mutant CTTNBP2. After nocodazole treatment, neurons were subjected to immunostaining using antibody against acetylated tubulin. Normalized signals of acetylated tubulin are shown. Data in (b), (d) and (e) represent mean ± SEM. In (d), each dot indicates an independent immunoblotting experiment. In (e), one dot represents the average result of one neuron. Data in (c) represent the data collected from three independent experiments. WT, n = 73, D570Y, n = 74. (b), (d) Two-way ANOVA; (e) two-tailed Mann–Whitney test. All statistical data and exact sample sizes are available in Additional file 2: Table S1. *P < 0.05; **P < 0.01; ***P < 0.001; ns, not significant. Scale bar: a 10 μm; e 5 μm