Fig. 5. CK1 and Wnt-5a mediate the changes in phosphorylation and cytoplasmic distribution of Dvl2. (A) The subcellular distribution of Dvl2-Myc in transfected SN4741 cells was assessed through an anti-Myc antibody by confocal microscopy. Four patterns were detected: (1) even distribution, and punctae of (2) small, (3) intermediate or, (4), large size. For a detailed description see the results section. (B) Co-transfection of Dvl2-Myc and CK1 or the kinase-dead CK1 (K>R) mutant resulted in a predominant even or punctate distribution of Dvl2-Myc, respectively. The arrow points to a cell transfected with Dvl2-Myc (Cy3, red) with no or low levels of exogenous CK1 (Cy2, green), serving as an internal control of the experiment. (C) Quantification of changes in the intracellular localization of Dvl2-Myc upon coexpression of CK1 or CK1 (K>R) as described in A and B. (D) D4476 (100 µM) blocks the phosphorylation dependent shift of Dvl2-Myc induced by different amounts (ng/well) of transfected CK1 in SN4741 cells. The position of dephosphorylated () and phosphorylated () Dvl2 is indicated. Data are representative of at least three independent replicates. (E-G) SN4741 cells were transfected with Dvl2-Myc and treated as indicated: (E) The general CK1 inhibitor D4476 (100 µM) and (F) the CK1/-specific inhibitor IC261 (20 µM) changed the subcellular localization of Dvl2-Myc to a predominant punctate distribution. This is in contrast with Wnt-5a treatment (100 ng/ml) (G), which leads to a more even distribution of Dvl2-Myc than in control. Data in C,E,F,G (n3) were statistically evaluated, Data represent the mean ± s.d., one-way ANOVA with Bonferroni post-tests (^*P<0.05, ^**P<0.01, ^***P<0.001).