FIGURE 3.

Overexpressing Zbed3 activates Wnt/β-catenin signaling by stabilizing cytoplasmic β-catenin. A, overexpression of Zbed3 activates LEF-1-dependent reporter gene. NIH3T3 cells were transfected with wild-type or mutant LEF-1 binding site reporter gene, respectively. The plasmid of LacZ was transfected as the negative control. One day later, transfected cells were treated with control or Wnt-3a conditioned medium (CM) for 6 h, and then luciferase activity was determined as described under “Experimental Procedures.” B, Zbed3 wild-type (WT) and mutants (SA or TA) exhibited different abilities in activating LEF-1-dependent reporter activities. LEF-1-dependent reporter plasmid was co-transfected with WT, SA, or TA forms of Zbed3 into NIH3T3 cells. 24 h later, cells were lysed for reporter assay. The expression of Zbed3 Wild-typed (WT), SA, and TA mutants are shown in the lower panel. C, Zbed3 could stabilize cytoplasmic β-catenin. Transfected NIH3T3 cells were stimulated with Wnt-3a CM for 3 h, and cytoplasmic fraction was collected and detected by anti-β-catenin antibody, and β-tubulin was used as cytoplasmic marker.