Fig. 8

DVL2 facilitates AURKB export from the nucleus and PLK4 stabilisation. a MDA-MB-231 cells were treated with control or DVL2 siRNAs for 72 h, and then stimulated overnight with DMEM or ACM. Cells were stained for actin, AURKB and nucleus (DAPI). Bar = 20 µm. b MDA-MB-231 cells were stimulated overnight with DMEM or ACM in the presence of DMSO or Leptomycin (LMB), a nuclear export inhibitor. Cells were stained with α-tubulin, AURKB and DAPI. Bar = 20 µm. c, d Quantification of the nuclear to cytoplasmic (N/C) ratio of AURKB from cells in (a) and (b), respectively. The data were plotted as described in Methods (***p < 0. 001; n = 3, at least 60 cells were measured per condition). e Cells were stimulated overnight with DMEM or ACM in the presence of DMSO, LMB or LMB + centrinone. Cell motility was measured as described in Methods (**p < 0.01, ***p < 0.001; n = 3, at least 40 cells were tracked per condition). f Model for DVL2 regulation of the nuclear and cytoplasmic/cortical pools of AURKB. g HEK293T cells were transfected with 3HA-PLK4, 3 Flag-BTRC and increasing amounts of T7-DVL2-wt or T7-DVL2 (Δ361–736) mutant. Co-immunoprecipitation was performed using anti-Flag antibody. h Quantification of the binding between PLK4 and BTRC in the presence of increasing amounts of DVL2 is shown as average signal ratio of HA-PLK4/3F-BTRC in immunoprecipitates. Data were analysed with two-way ANOVA post-tested with Bonferroni test (n = 4, error bars are ± s.e.m.). i Model for DVL2-mediated PLK4 stabilisation. The data from c, d and e were plotted as box and whiskers. Boxes represent median and 25th to 75th percentiles, whiskers the minimum and maximum values with each individual cell value superimposed. Data were compared with one-way ANOVA Kruskal–Wallis test and post-tested with Dunn’s multiple comparison test. Arrow indicates the control bar used for comparison