Upon binding of a Wnt ligand to the frizzled (FZD)-low density lipoprotein receptor related protein 5/6 (LRP5/6) receptor complex, the beta-catenin destruction complex, composed of Axinl, adenomatous polyposis coli (APC), glycogen synthase kinase 3 (GSK3) and casein kinase 1 (CK1), is immediately inactivated, which causes beta-catenin stabilization. However, the molecular mechanism of signal transduction from the receptor complex to the beta-catenin destruction complex is controversial. Here we show that Wnt3a treatment promotes the dissociation of the Axinl-APC complex in glioblastoma cells cultured in serum-free medium. Experiments with the GSK3 inhibitor BIO suggest that Axinl -APC dissociation was controlled by phosphorylation. Introduction of a phosphomimetic mutation into Thr160 of Axinl, located in the APC-binding region RGS, abrogated the interaction of Axinl with APC. Consistent with these observations, the Axinl phosphomimetic mutant lost the ability to reduce beta-catenin stability and to repress beta-catenin/TCF-dependent transcription. Taken together, our results suggest a novel mechanism of Wnt signaling through the dissociation of the beta-catenin destruction complex by Axinl Thr160 modification. (C) 2015 Elsevier Inc. All rights reserved.