Figure 2

Down-regulation of CDK5 reduces CRMP2 phosphorylation in NPC-derived neural progeny. Differentiating NPCs were treated on day 2 with the pharmacological inhibitor Roscovitine (Rosco) or siRNA against CDK5 (siCDK5), followed by infection with p35-adenovirus. On day 4, NPC-derived neural progeny were fixed for immunofluorescence or lysed for immunoblot analysis. (A-C) Immunocytochemical analysis with an antibody against pSer522-CRMP2 showed low levels of endogenous immunoreactivity in uninfected, vehicle-treated cells (A), with a slight reduction in cells treated with Roscovitine (B) or siCDK5 (C). (D-F) NPC-derived neural progeny expressing p35 showed a notable increase in CRMP2 phosphorylation (D) that was reversed by treatment with Rosco (E) or siCDK5 (F). (G) Reversal of CRMP2 hyperphosphorylation with CDK5 down-regulation. (H) Immunoblot analysis of pSer522-CRMP2, total (t)CRMP2, CDK5, and actin as a loading control. (I) Semi-quantitative image analysis of levels of CRMP2 phosphorylation in NPCs infected with p35-adv with or without Rosco or siCDK5. * p < 0.05 compared to vehicle-treated controls by one-way ANOVA with post-hoc Dunnett's test. # p < 0.05 compared to p35-expressing NPCs by one-way ANOVA with post-hoc Tukey-Kramer test. Scale bar = 10 μm.