Fig. 5. mDia negatively regulates -catenin localization. (A) The specific GSK3 inhibitor BIO induces nuclear localization of endogenous -catenin (red) and increased cell-cell contact (phase) in myoblasts after a 24-hour treatment in growth medium. Bar, 10 µm (25 µm in phase). (B) mDiaN3 inhibits -catenin nuclear localization in BIO-treated cells. Myoblasts were transiently transfected with control (EGFP), mDia1-FL, N3, HIND3, H+P or F2 constructs (all GFP-tagged), treated with 2.5 µM BIO for 24 hours and stained for -catenin (red). In N3 and HIND3-transfected cells, accumulation of -catenin at cell-cell contacts correlates with loss of nuclear staining. (C) Confocal analysis shows absence of -catenin staining in the nuclei of N3-transfected cells. Bar, 10 µm. (D) Quantification of the effects of GFP-tagged mDia constructs on -catenin localization (mean ± s.e.m., n=2).