Fig. 5

Aβ and TTBK1 induce CRMP2 phosphorylation in vitro. a HEK293 cells were transfected with CRMP2 expression plasmids [wild type (WT), S522A mutant, and T555A mutant] with or without human wild type TTBK1 plasmid, followed by immunoblotting of total and phospho-CRMP2 at T514. b Schematic diagram of site-specific CRMP2 phosphorylation by TTBK1 and Aβ42. c-d SH-SY5Y cells were transfected with scramble or TTBK1 siRNA, followed by stimulation with different doses of freshly prepared Aβ42 (0, 3, and 10 μM) for 24 h. Cell lysates were subjected to immunoblotting of endogenous pCRMP2 at T514, and endogenous TTBK1. e SH-SY5Y cells were transfected with a combination of TTBK1 and DN-Rho plasmids (0.5 μg each), followed by stimulation with 10 μM freshly prepared Aβ42 for 24 h. Results were representative of 3 independent experiments. The error bars indicate SEM. * and ** denote p < 0.05, or 0.01 as determined by one-way ANOVA and Tukey post hoc for C and E, and by Student’s t-test for D (N = 3 per group). C, ANOVA F (2, 9) = 34.8591 (p < 0.0001), E, ANOVA F (3, 8) = 36.2727 (p < 0.0001)