Negative feedback regulation of Wnt signaling by Gβγ-mediated reduction of Dishevelled

Wnt signaling is known to be important for diverse embryonic and post-natal cellular events and be regulated by the proteins Dishevelled and Axin. Although Dishevelled is activated by Wnt and involved in signal transduction, it is not clear how Dishevelled-mediated signaling is turned off. We report that guanine nucleotide binding protein beta 2 (Gnb2; Gβ₂) bound to Axin and Gβ₂ inhibited Wnt mediated reporter activity. The inhibition involved reduction of the level of Dishevelled, and the Gβ ₂γ₂ mediated reduction of Dishevelled was countered by increased expression of Axin. Consistent with these effects in HEK293T cells, injection of Gβ₂γ₂ into Xenopus embryos inhibited the formation of secondary axes induced either by XWnt8 or Dishevelled, but not by β-catenin. The DEP domain of Dishevelled is necessary for both interaction with Gβ₂γ₂ and subsequent degradation of Dishevelled via the lysosomal pathway. Signaling induced by Gβ₂γ₂ is required because a mutant of Gβ₂, Gβ₂ (W332A) with lower signaling activity, had reduced ability to downregulate the level of Dishevelled. Activation of Wnt signaling by either of two methods, increased Frizzled signaling or transient transfection of Wnt, also led to increased degradation of Dishevelled and the induced Dishevelled loss is dependent on Gβ₁ and Gβ₂. Other studies with agents that interfere with PLC action and calcium signaling suggested that loss of Dishevelled is mediated through the following pathway: Wnt/Frizzled →Gβγ→ PLC →Ca⁺²/PKC signaling. Together the evidence suggests a novel negative feedback mechanism in which Gβ₂γ₂ inhibits Wnt signaling by degradation of Dishevelled.