Schneider PN, Slusarski DC, Houston DW. (2012) Differential role of axin RGS domain function in Wnt signaling during anteroposterior patterning and maternal axis formation. PLoS One, 7(9):e44096.
Axin is part of a multi-protein complex that facilitates the phosphorylation and degradation of β-catenin, a critical transcription regulator involved in development. In the presence of Wnt signaling, axin is removed from its role in the β-catenin destruction complex, and β-catenin is able to enter the nucleus to activate gene transcription. In this paper, the authors investigate an RGS (Regulator of G-protein Signaling) domain within axin to see if it has a larger role in regulating activity of the β-catenin destruction complex. The authors do this by mutating the RGS domain of axin, while leaving the other adenomatous polyposis coli (APC, a protein that helps regulate how cells divide) interactions and degradation functions intact. The mutant is studied in vitro
and in vivo
using Xenopus laevis
and Danio rerio
. To confirm that the axin mutant is still able to interact with APC and the destruction complex, the authors cloned a 459 bp portion of the Xenopus
APC producing the axin-interacting region. The authors were able to easily generate the control Xenopus
APC fragment by ordering it as a single gBlocks® Gene Fragment
and cloning it into a plasmid containing an HA tag sequence. The expressed fragment was then used to confirm that the wild type and mutant axin RGS domains were able to interact normally with APC.