RAMP and MRAP Accessory Proteins have Selective Effects on Expression and Signalling of the CB1, CB2, GPR18 and GPR55 Cannabinoid Receptors

Background and Purpose Receptor activity‐modifying proteins (RAMPs) and melanocortin receptor accessory proteins (MRAPs) modulate expression and signalling of calcitonin and melanocortin GPCRs. Interactions with other GPCRs have also been reported. The cannabinoid receptors, CB 1 and CB 2 , and two putative cannabinoid receptors, GPR18 and GPR55, exhibit substantial intracellular expression and there are discrepancies in ligand responsiveness between studies. We investigated whether interactions with RAMPs or MRAPs could explain these phenomena. Experimental Approach Receptors and accessory proteins were co‐expressed in HEK‐293 cells. Selected receptors were studied at basal expression levels and also with enhanced expression produced by incorporation of a preprolactin signal sequence/peptide (pplss). Cell surface and total expression of receptors and accessory proteins were quantified using immunocytochemistry. Signalling was measured using cAMP (CAMYEL) and G protein dissociation (TRUPATH Gα 13 ) biosensors. Key Results MRAP2 enhanced surface and total expression of GPR18. Pplss‐GPR18 increased detection of cell surface MRAP2. MRAP1α and MRAP2 reduced GPR55 surface and total expression, correlating with reduced constitutive, but not agonist‐induced, signalling. GPR55, pplss‐CB 1 and CB 2 reduced detection of MRAP1α at the cell surface. Pplss‐CB 1 agonist potency was reduced by MRAP2 in Gα 13 but not cAMP assays, consistent with MRAP2 reducing pplss‐CB 1 expression. Some cannabinoid receptors increased RAMP2 or RAMP3 total expression without influencing surface expression. Conclusions and Implications Mutual influences on expression and/or function for specific accessory protein‐receptor pairings raises the strong potential for physiological and disease‐relevant consequences. Sequestration and/or hetero‐oligomerisation of cannabinoid receptors with accessory proteins is a possible novel mechanism for receptor crosstalk.