Rapid, Wnt-Induced Changes in GSK3β Associations that Regulate β-Catenin Stabilization Are Mediated by Gα Proteins

Background: In the absence of Wnt stimulation, the transcriptional cofactor b-catenin is destabilized via phosphorylation by protein kinase GSK3b in complex with Axin family members. In the ‘‘canonical’’ Wnt signaling pathway, Disheveled (Dvl) is required to functionally inhibit the activity of the GSK3b/Axin complex and thereby stabilize b-catenin. Yet, the mechanisms that underlie Wnt regulation of GSK3 and stabilization of b-catenin are still not fully appreciated. Results: Here, we examine time-dependent changes in protein-protein interactions that occur in response to Wnt treatment. We show that GSK3b/Axin complexes are rapidly (t1/2 < 3 min) disrupted upon Wnt stimulation and that changes in GSK3b/Axin association substantially precede both b-catenin stabilization and Axin degradation. We further demonstrate that depletion of Gao or Gaq will inhibit, respectively, the Wnt-induced disruption of GSK3b/Axin2 and GSK3b/Axin complexes and diminish Wnt stabilization of b-catenin. We also show that direct activation of G proteins in vivo with GTPgS in the absence of exogenous Wnt will disrupt GSK3b/Axin2 complexes and stabilize b-catenin. Finally, we demonstrate an association of Gao with Fz that is also very rapidly (t1/2 < 1 min) perturbed upon Wnt-3a stimulation and that the Wnt-dependent effects on both GSK3b/Axin2 and Gao/Fz are pertussis-toxin sensitive. Collectively, these data implicate a role for G proteins in the regulation of Wnt-mediated protein-protein interactions and signaling to b-catenin. Conclusions: We conclude that rapid disruption of GSK3b/Axin interactions in response to Wnt leads to the initial stabilization of b-catenin and that Gao and Gaq signaling contributes to Wnt-mediated GSK3b/Axin disruption and the ultimate stabilization of b-catenin.