RAMP2 influences glucagon receptor pharmacology via trafficking and signaling

Endogenous satiety hormones provide an attractive target for obesity drugs. Glucagon causes weight loss by reducing food intake and increasing energy expenditure. To further understand the cellular mechanisms by which glucagon and related ligands activate the glucagon receptor (GCGR), we have investigated the interaction of the GCGR with RAMP2, a member of the family of Receptor Activity Modifying Proteins. We have used a combination of competition binding experiments, cell surface ELISA, functional assays assessing the Gαs and Gq pathways and β-arrestin recruitment, and siRNA knockdown to examine the effect of RAMP2 on the GCGR. Ligands tested were glucagon, glucagon-like peptide-1(GLP-1), oxyntomodulin and analogue G(X), a GLP-1/glucagon co-agonist developed in-house.
Confocal microscopy was employed to assess whether RAMP2 affects the subcellular distribution of GCGR. Here we demonstrate that co-expression of RAMP2 and the GCGR results in reduced cell surface expression of the GCGR. This was confirmed by confocal microscopy, which demonstrated that RAMP2 co-localises with the GCGR and causes significant GCGR cellular redistribution.
Furthermore, the presence of RAMP2 influences signalling through the Gαs and Gαq pathways, as well as recruitment of β-arrestin. This work suggests that RAMP2 may modify the agonist activity and trafficking of the GCGR, with potential relevance to production of new peptide analogues with selective agonist activities.