Title: Rapid, Wnt-Induced Changes in GSK3β Associations that Regulate β-Catenin Stabilization Are Mediated by Gα Proteins
Abstract: BackgroundIn the absence of Wnt stimulation, the transcriptional cofactor β-catenin is destabilized via phosphorylation by protein kinase GSK3β in complex with Axin family members. In the “canonical” Wnt signaling pathway, Disheveled (Dvl) is required to functionally inhibit the activity of the GSK3β/Axin complex and thereby stabilize β-catenin. Yet, the mechanisms that underlie Wnt regulation of GSK3 and stabilization of β-catenin are still not fully appreciated.ResultsHere, we examine time-dependent changes in protein-protein interactions that occur in response to Wnt treatment. We show that GSK3β/Axin complexes are rapidly (t1/2 < 3 min) disrupted upon Wnt stimulation and that changes in GSK3β/Axin association substantially precede both β-catenin stabilization and Axin degradation. We further demonstrate that depletion of Gαo or Gαq will inhibit, respectively, the Wnt-induced disruption of GSK3β/Axin2 and GSK3β/Axin complexes and diminish Wnt stabilization of β-catenin. We also show that direct activation of G proteins in vivo with GTPγS in the absence of exogenous Wnt will disrupt GSK3β/Axin2 complexes and stabilize β-catenin. Finally, we demonstrate an association of Gαo with Fz that is also very rapidly (t1/2 < 1 min) perturbed upon Wnt-3a stimulation and that the Wnt-dependent effects on both GSK3β/Axin2 and Gαo/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 β-catenin.ConclusionsWe conclude that rapid disruption of GSK3β/Axin interactions in response to Wnt leads to the initial stabilization of β-catenin and that Gαo and Gαq signaling contributes to Wnt-mediated GSK3β/Axin disruption and the ultimate stabilization of β-catenin.