Phosphorylation of Axin, a Wnt signal negative regulator, by glycogen synthase kinase-3 beta regulates its stability

Axin forms a complex with glycogen synthase kinase-3 beta (GSK-3 beta) and beta-catenin and promotes GSK-3 beta-dependent phosphorylation of beta-cate nin, thereby stimulating the degradation of beta-catenin. Because GSH-3 bet a also phosphorylates Axin in the complex, the physiological significance o f the phosphorylation of Axin was examined. Treatment of COS cells with LiC l, a GSK-3 beta inhibitor, and okadaic acid, a protein phosphatase inhibito r, decreased and increased, respectively, the cellular protein level of Axi n. Pulse-chase analyses showed that the phosphorylated form of Axin was mor e stable than the unphosphorylated form and that an Axin mutant, in which t he possible phosphorylation sites for GSK-3 beta were mutated, exhibited a shorter half-life than wild type Axin, Dvl-1, which was genetically shown t o function upstream of GSK-3 beta, inhibited the phosphorylation of Axin by GSK-3 beta in vitro. Furthermore, Wnt-3a-containing conditioned medium dow n-regulated Axin and accumulated beta-catenin in L cells and expression of DV1-1(Delta PDZ), in which the PDZ domain was deleted, suppressed this acti on of Wnt-3a, These results suggest that the phosphorylation of Axin is imp ortant for the regulation of its stability and that Wnt down-regulates Axin through Dv1.