Identification of MRAP protein family as broad‐spectrum GPCR modulators

Abstract

Background The melanocortin receptor accessory proteins (MRAP1 and MRAP2) are well‐known endocrine regulators for the trafficking and signalling of all five melanocortin receptors (MC1R–MC5R). The observation of MRAP2 on regulating several non‐melanocortin G protein‐coupled receptors (GPCRs) has been sporadically reported, whereas other endogenous GPCR partners of the MRAP protein family are largely unknown. Methods Here, we performed single‐cell transcriptome analysis and drew a fine GPCR blueprint and MRAPs‐associated network of two major endocrine organs, the hypothalamus and adrenal gland at single‐cell resolution. We also integrated multiple bulk RNA‐seq profiles and single‐cell datasets of human and mouse tissues, and narrowed down a list of 48 GPCRs with strong endogenous co‐expression correlation with MRAPs. Results 36 and 46 metabolic‐related GPCRs were consequently identified as novel interacting partners of MRAP1 or MRAP2, respectively. MRAPs exhibited protein–protein interactions and varying pharmacological properties on the surface translocation, constitutive activities and ligand‐stimulated downstream signalling of these GPCRs. Knockdown of MRAP2 expression by hypothalamic administration of adeno‐associated virus (AAV) packed shRNA stimulated body weight gain in mouse model. Co‐injection of corticotropinreleasing factor (CRF), the agonist of corticotropin releasing hormone receptor 1 (CRHR1), suppressed feeding behaviour in a MRAP2‐dependent manner. Conclusions Collectively, our study has comprehensively elucidated the complex GPCR networks in two major endocrine organs and redefined the MRAP protein family as broad‐spectrum GPCR modulators. MRAP proteins not only serve as a vital endocrine pivot on the regulation of global GPCR activities in vivo that could explain the composite physiological phenotypes of the MRAP2 null murine model but also provide us with new insights of the phenotyping investigation of GPCR–MRAP functional complexes.

Single transmembrane GPCR modulating proteins: neither single nor simple

“…We performed a protein alignment analysis and found that the S19 and Y250 in MC2R were not conserved in the newly identified GPCRs that bind to MRAP1 from our study ( Fig. 2 ) ( Wang et al, 2022 a). This is probably because MRAP1 needs this interface to regulate MC2R activity but not for direct interaction.…”

“…Benefiting from the rapid development of multi-omics technologies, we first acquired numerous metabolic-related GPCRs that coexpressed with MRAP2 in the same neurons via single-cell RNA sequencing (scRNA-seq). Next, we experimentally screened the majority of GPCRs and found that they not only exhibited direct interactions with MRAP2 proteins but also pharmacologically regulated by MRAP2 in terms of cell surface expression and ligand-induced downstream cascades ( Wang et al, 2022 a). This work redefined MRAP proteins as broad-spectrum modulators of metabolic-related GPCR signaling in vitro and in vivo ( Fig.…”

MRAP2 Inhibits β-Arrestin-2 Recruitment to the Prokineticin Receptor 2

“…This type of regulator binds an allosteric site that is distinct from the orthosteric site recognised by endogenous ligands and stabilises a specific receptor conformation. The characterisation of the natural allosteric modulation of GPCRs is a promising new approach for GPCR drug discovery compared to classical approaches that focus on the development of small molecules targeting the orthosteric site [51][52][53][54][55][56].…”

MC4R in Central and Peripheral Systems

“…MRAP exhibits protein‐protein interactions and distinct pharmacological properties on surface translocation, homeodomain activity, and ligand‐stimulated downstream signaling of these GPCRs. [ 122 ] Prokinetic peptides (PKs) are agonists of G protein‐coupled receptors: prokinetic receptors (PKRs), and they reduce food intake in rodents. The inhibition of food intake by PKs is mediated through amygdala PKR2 neurons.…”