Analyzing the signaling properties of gar (Lepisosteus oculatus) melanocortin receptors: Evaluating interactions with MRAP1 and MRAP2

•Gar Mc2r requires co-expression with gar Mrap1 for activation in CHO cells.•Gar Mc1R and Gar Mc3r are more sensitive to activation by NDP-MSH than to hACTH(1–24).•Gar Mc4r, and Mc5r can be activated equally well by either NDP-MSH and hACTH(1–24).•Co-expression of gar Mc4r with Mrap2 increases Vmax following stimulation with ACTH(1–24) or NDP-MSH.•Co-expression of gar Mc5r with gar Mrap1 increases trafficking of the receptor to the plasma membrane. RT-PCR analysis of gar pituitary and brain indicated that different combinations of gar melanocortin receptor mRNAs are present in the same tissues with mRNAs for gar mrap1 and gar mrap2. Against this background, an objective of this study was to determine whether the ligand sensitivity for either ACTH or α-MSH was affected when gar (g) melanocortin receptors (Mcrs) were co-expressed with either of the accessory proteins gMrap1 or gMrap2 in Chinese Hamster Ovary cells. The results indicated that gMc2r has an obligatory requirement for co-expression with gMrap1 in order for the receptor to be activated by hACTH(1–24). In addition, activation of gMc2r did not occur when the receptor was expressed alone or co-expressed with gMrap2. Furthermore, co-expression of gMc2r with gMrap1 followed by stimulation with NDP-MSH resulted in a low level of activation (only at 10−7 M and 10−6 M). However, gMc1r, gMc3r, gMc4r, and gMc5r responded to stimulation by NDP-MSH in a more robust manner. Co-expression of gMc1r, gMc3r, gMc4r, and gMc5r with gMRAP1 had no effect on sensitivity to stimulation by NDP-MSH or hACTH(1–24). Co-expression with gMRAP2 had no negative or positive effect on ligand sensitivity for gMc1r, gMc3r, and gMc5r, however this treatment did increase the activation of CHO cells transfected with gMc4r following stimulation with both hACTH(1–24) (p