Fig. 2. CK1 inhibition blocks Wnt-5a-induced phosphorylation of Dvl2 and Dvl3. SN4741 cells were treated with vehicle (control) or Wnt-5a (100 ng/ml) for 2 hours in the presence or absence of D4476 (100 µM) (A) or IC261 (10 µM) (C). Western blot analysis was performed as in Fig. 1. The position of dephosphorylated () and phosphorylated () Dvl is indicated. Antibodies against phospho-Ser45--catenin (CK1 target site), against active (Ser37-Thr41-dephosphorylated) -catenin (ABC) and total -catenin were used. (B) Scheme of phosphorylation sites of -catenin. Ser45 is a CK1 target site, which serves as a priming for sequential phosphorylation of Thr41, Ser37 and Ser33 by GSK3. (D) SN4741 cells were transfected with plasmids encoding Dvl2-Myc and either CK1 or kinase-dead CK1 (K>R) mutant. Phosphorylation of Dvl2-Myc was detected as a phosphorylation-dependent mobility shift by Myc- and Dvl2-specific antibodies. CK1 levels were monitored with a CK1 specific antibody. Data in A, C and D are representative of at least three independent replicates.