Structural and biochemical characterization of the axin/adenomatous polyposis coli/beta-catenin Wnt signaling complex

The Wnt growth factor signaling pathway plays an essential role in the development of diverse organisms from Dictyostelium to humans. Inappropriate activation of Wnt signaling by mutation of pathway components has been observed in a vast number of human cancers. In the absence of a Wnt signal, the tumor suppressors Axin and Adenomatous Polyposis Coli (APC) directly interact with and serve to regulate cytoplasmic levels of the proto-oncogene β-catenin. Both Axin and APC are known to be critical for β-catenin regulation, and truncations in APC that eliminate Axin and β-catenin binding sites result in human cancers. The region of Axin responsible for APC binding is a member of the regulator of G-protein signaling (RGS) superfamily. β-catenin binding by APC occurs through a series of 15- and 20-amino acid repeat sequences. Likewise, APC binds to Axin using characteristic sequences known as ‘SAMP repeats’.; We have used structural and biochemical methods to study APC and its interactions with Axin and β-catenin. Circular dichroism spectra and other biochemical data indicate that the region of APC responsible for β-catenin and Axin binding is unstructured in the absence of these proteins. Gel-filtration chromatography was used to measure the stoichiometry of the APC/β-catenin interaction. The x-ray crystal structure of the RGS domain of Axin in complex with a SAMP repeat from APC reveals that the Axin-APC interaction occurs at a conserved groove on a face of the RGS domain that is distinct from the G-protein interface of classical RGS proteins. The region of Axin-RGS corresponding to the classical RGS-G-protein interface is not conserved in Axin-family RGS domains. This suggests that, despite their homology, Axin-family and classical RGS proteins have distinct functions.; The crystal structure of a β-catenin/APC 15-mer complex reveals unique features of the APC/β-catenin interaction not seen in previous β-catenin/ligand complex structures, suggesting a potential binding site for therapeutics. Based on the 15-mer complex structure, we propose a model for 20-mer binding in which the 20-mers adopt alternate modes of binding to β-catenin with a switch between modes triggered by phosphorylation.