Crosstalk between G proteins and the canonical Wnt signaling pathway

Signal transduction through heterotrimeric G proteins results in a wide variety of cellular responses initiated by an interaction between G proteins and G protein-coupled receptors. Receptor-driven activation of G proteins induces the dissociation of the trimeric G protein complex and gives rise to two signaling-competent species: GTP-Galpha and a Gbetagamma dimer, which are both able to interact with downstream effectors to trigger signaling cascades. Signaling is terminated when GTP (bound to Galpha) is hydrolyzed back to GDP, and the heterotrimeric complex is re-established. GTP hydrolysis is often significantly accelerated by members of a family of proteins called R&barbelow;egulators of G&barbelow;-protein S&barbelow;ignaling (RGS proteins) that bind preferentially to the active Galpha.; Recent observations have suggested the potential involvement of G-proteins in Wnt signaling, a pathway that is important in diverse developmental processes and can be involved in tumor formation. The primary role of the so-called canonical Wnt pathway is to regulate cytoplasmic levels of beta-catenin, a multi-functional protein that can act as a transcriptional co-activator when it is permitted to accumulate. Interestingly, the major negative regulator of canonical Wnt signaling, axin, contains the conserved domain common to all RGS proteins. Despite its sequential and structural homology to the RGS family, no activity had yet been determined for this region of axin with respect to G-proteins. I have uncovered a specific, activation-dependent interaction between the RGS domain of axin and the alpha subunit of the heterotrimeric G protein G12. While binding to axin had no detectable effect on the GTP hydrolysis activity of Galpha12, in cell-based studies this interaction did contribute to the negative regulation of canonical Wnt signaling. In contrast, the closely related G12 family member Galpha13 did not bind axin and could not negatively regulate Wnt signaling. Quite surprisingly, however, activated Galpha13 had a significant stimulatory effect on the Wnt pathway.; Further analysis of the stimulatory effect of Galpha13 on beta-catenin-dependent signaling demonstrated that this activity was due to the Galpha13-mediated activation of RhoA, a monomeric G protein. I found that expression of activated RhoA in 293T cells induced upregulation of TCF-mediated transcriptional events in a beta-catenin and TCF-dependent manner. Furthermore, inhibition of RhoA by Clostridium botulinum toxin C3 attenuated Wnt3A-stimulated signaling through the canonical pathway by at least 50%, demonstrating that RhoA has a significant role in canonical Wnt signaling. RhoA does not appear to effect the stabilization or nuclear accumulation of beta-catenin, which is reported to be the primary means by which beta-catenin dependent transcription occurs. While the mechanism by which RhoA activates beta-catenin dependent transcription remains to be elucidated, the requirement for RhoA in the Wnt pathway is an exciting discovery that may reshape our understanding of the canonical Wnt signaling cascade.