Rapid, Wnt-induced changes in GSK3beta associations that regulate beta-catenin stabilization are mediated by Galpha proteins.
BACKGROUND
In the absence of Wnt stimulation, the transcriptional cofactor beta-catenin is destabilized via phosphorylation by protein kinase GSK3beta in complex with Axin family members. In the "canonical" Wnt signaling pathway, Disheveled (Dvl) is required to functionally inhibit the activity of the GSK3beta/Axin complex and thereby stabilize beta-catenin. Yet, the mechanisms that underlie Wnt regulation of GSK3 and stabilization of beta-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 GSK3beta/Axin complexes are rapidly (t1/2 < 3 min) disrupted upon Wnt stimulation and that changes in GSK3beta/Axin association substantially precede both beta-catenin stabilization and Axin degradation. We further demonstrate that depletion of Galpha(o) or Galpha(q) will inhibit, respectively, the Wnt-induced disruption of GSK3beta/Axin2 and GSK3beta/Axin complexes and diminish Wnt stabilization of beta-catenin. We also show that direct activation of G proteins in vivo with GTPgammaS in the absence of exogenous Wnt will disrupt GSK3beta/Axin2 complexes and stabilize beta-catenin. Finally, we demonstrate an association of Galpha(o) with Fz that is also very rapidly (t1/2 < 1 min) perturbed upon Wnt-3a stimulation and that the Wnt-dependent effects on both GSK3beta/Axin2 and Galpha(o)/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 beta-catenin.


CONCLUSIONS
We conclude that rapid disruption of GSK3beta/Axin interactions in response to Wnt leads to the initial stabilization of beta-catenin and that Galpha(o) and Galpha(q) signaling contributes to Wnt-mediated GSK3beta/Axin disruption and the ultimate stabilization of beta-catenin.
Liu
Xunxian
X
Laboratory of Cellular and Developmental Biology, NIDDK, National Institutes of Health, Bethesda, Maryland 20892, USA.
Rubin
Jeffrey S
JS
Kimmel
Alan R
AR
D000818:Animals; D015153:Blotting, Western; D002460:Cell Line; D003598:Cytoskeletal Proteins; D044385:GTP-Binding Protein alpha Subunits / Q000378:metabolism*; D019204:GTP-Binding Proteins / Q000378:metabolism*; D018507:Gene Expression Regulation, Developmental / Q000502:physiology*; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D047468:Immunoprecipitation; D051379:Mice; D009687:Nuclear Proteins; D010766:Phosphorylation; D034741:RNA, Small Interfering / Q000235:genetics; D015398:Signal Transduction / Q000502:physiology*; D010738:Type C Phospholipases; D051153:Wnt Proteins / Q000378:metabolism*; D051176:beta Catenin / Q000378:metabolism*
C495267:CTNNB1 protein, mouse; D003598:Cytoskeletal Proteins; D044385:GTP-Binding Protein alpha Subunits; C099035:Nin protein, mouse; D009687:Nuclear Proteins; D034741:RNA, Small Interfering; D051153:Wnt Proteins; D051176:beta Catenin; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D010738:Type C Phospholipases; D019204:GTP-Binding Proteins