Figure 4. Changes in expression and phosphorylation of the proteins in cancer cells following GSK3β inhibition.

(A) Immunoblotting analysis compares the expression of Rb, CDK4, CDK6 and cyclin D1, and the phosphorylation of Rb at S780 and S807/811 residues (p-Rb^S780, p-Rb^S807/811) between cells treated with DMSO (DM) or 10 µM AR-A014418 (AR) for 24 hrs. (B) Changes in levels of p-Rb^S780 and p-Rb^S807/811 and expression of Rb and cyclin D1 were examined in MIA PaCa-2 cells at the indicated time points after treatment with 10 µM AR-A014418. (C) Expression of Rb, cyclin D1, GSK3α and GSK3β proteins and levels of Rb phosphorylation (p-Rb^S780) were examined and compared between the same pancreatic cancer cells transfected with non-specific siRNA (NS) or GSK3β-specific siRNA (S) (10 nM each). (A–C) β-actin expression was monitored as a loading control.