Effect of HDAC6 inhibitor-mediated
increase in acetylation of β-catenin
Lys49 on N-terminal posttranslational modification sites. (A) Time-course
immunoblot analysis of human NPCs treated with an HDAC6 inhibitor.
Human NPCs were treated with HDAC6 inhibitor ACY-1215 at 5 μM
for the times points indicated and the lysates immunoblotted with
antibodies against β-catenin, Ac-Lys49-β-catenin, phos-Ser45
and phos-Ser33, Ser37, and Thr41. β-actin is using shown as
loading control. (B) Quantification of the intensity of Western blot
band signals from A. in indicated antigens normalized
to total β-catenin before and after treatment with ACY-1215
at 5 μM for 8 h. Results are representative of three independent
experiments. Error bars indicate standard deviation. ** and *** denote
significance with p < 0.01 and p < 0.001, respectively (paired t test). (C) Immunofluorescence
staining for HDAC6 in human NPCs when treated with control siRNA (left)
and HDAC6 siRNA (right). (D) Immunoblot analysis of lysates from human
NPCs treated with HDAC6 siRNA, control siRNA, and ACY-1215, using
antibodies against HDAC6, β-catenin, Ac-Lys49-β-catenin,
and phos-Ser45-β-catenin. β-actin is shown as loading
control. (E) Time course of CK1α and GSK3β bound to β-catenin
bound to in human NPCs treated with 5 μM ACY-1215. Cell lysates
were immunoprecipitated with anti-β-catenin antibody and levels
of CK1α and GSK3β were measured by immunoblot analysis.