Glycogen synthase kinase‐3 (GSK‐3) mediates epidermal growth factor, insulin and Wnt signals to various downstream events such as glycogen metabolism, gene expression, proliferation and differentiation. We have isolated here a GSK‐3β‐interacting protein from a rat brain cDNA library using a yeast two‐hybrid method. This protein consists of 832 amino acids and possesses Regulators of G protein Signaling (RGS) and dishevelled (Dsh) homologous domains in its N‐and C‐terminal regions, respectively. The predicted amino acid sequence of this GSK‐3β‐interacting protein shows 94% identity with mouse Axin, which recently has been identified as a negative regulator of the Wnt signaling pathway; therefore, we termed this protein rAxin (r at Axin). rAxin interacted directly with, and was phosphorylated by, GSK‐3β. rAxin also interacted directly with the armadillo repeats of β‐catenin. The binding site of rAxin for GSK‐3β was distinct from the β‐catenin‐binding site, and these three proteins formed a ternary complex. Furthermore, rAxin promoted GSK‐3β‐dependent phosphorylation of β‐catenin. These results suggest that rAxin negatively regulates the Wnt signaling pathway by interacting with GSK‐3β and β‐catenin and mediating the signal from GSK‐3β to β‐catenin.