Currently, the A-class G protein-coupled receptor 83 (GPR83) is officially classified as an orphan receptor. It is widely present in vertebrates and is highly conserved from fishes to mammals. GPR83 is primarily expressed in the brain, and is involved in the regulation of energy metabolism and certain anxiety-related behaviors (Dubins et al. 2012; Fakira et al. 2019, 2021; Müller et al. 2013). The human GPR83 gene can produce two transcripts (NM_016540 and NM_001330345) that either encode longer isoform 1 with seven transmembrane domains (TMDs) or shorter isoform 2 lacking TMD3. In previous publications and in the present study, GPR83 refers to the longer isoform 1 unless otherwise stated.
In recent years, the proprotein convertase subtilisin/kexin type 1 inhibitor (PCSK1N, also known as proSAAS)-derived peptide PEN (Gomes et al. 2016), the procholecystokinin (proCCK)-derived peptide proCCK56-63 (Mack et al. 2022), and family with sequence similarity 237 member A (FAM237A) (Sallee et al. 2020) were all reported as efficient agonists for GPR83. In a recent study (Li et al. 2023), we confirmed that human FAM237A binds to human GPR83 with nanomolar range affinity and activates it with nanomolar range efficiency using the NanoLuc Binary Technology (NanoBiT)-based ligand−receptor binding assay, fluorescent ligand-based visualization, and the NanoBiT-based β-arrestin recruitment assay; however, we did not detect any interaction between peptide PEN and proCCK56-63 with this receptor using these assays. Thus, it seemed that FAM237A, rather than peptide PEN and proCCK56-63, is the ligand for GPR83.
FAM237A was also named neurosecretory protein GL (NPGL) by Ukena's group (Ukena et al. 2014), and its biological function has been studied by this group in recent years (Fukumura et al. 2021a, 2021b, 2021c; Iwakoshi-Ukena et al. 2017; Matsuura et al. 2017; Narimatsu et al. 2021, 2022a, 2022b, 2022c; Shikano et al. 2018a, 2019, 2020). Administration or overexpression of FAM237A in murine or avian models typically increased food intake and fat accumulation, suggesting this neuropeptide is implicated in the regulation of energy metabolism. Consistently, knockout of Gpr83 in a mouse model led to a leaner phenotype with less fat accumulation (Müller et al. 2013). Thus, it seemed that FAM237A is an endogenous agonist for receptor GPR83.
FAM237B (named NPGM by Ukena's group) is a paralog of FAM237A. FAM237B is also widely present from fish to mammals and is conserved in evolution (Fig. S1). Both FAM237B and FAM237A are synthesized in vivo as precursors, including an N-terminal signal peptide, a mature peptide, and a C-terminal propeptide. At the conjunction of their predicted mature peptide and propeptide, a Gly residue and a dibasic cleavage site are absolutely conserved (Fig. S1), implying C-terminal α-amidation of their mature peptide. Moreover, both proteins contain two absolutely conserved Cys residues in their predicted mature peptide (Fig. S1), implying the formation of an intramolecular disulfide bond. Preliminary studies suggested that the biological function of FAM237B is similar to that of FAM237A (Kato et al. 2021; Martinez et al. 2023; Shikano et al. 2018b).
Mature human FAM237B and FAM237A share considerable sequence similarity, especially at their C-terminal fragment (Fig. 1a). Predicted by the AlphaFold algorithm, they also displayed similar three-dimensional structures, with their C-terminal fragment forming a long α-helix (Fig. 1b). A previous study reported that FAM237B is a weak agonist for GPR83: as high as 10 nM of FAM237B had no detectable activation effect in an inositol 1-phosphate accumulation assay (Sallee et al. 2020). In the present study, we prepared mature human FAM237B via an intein-fusion approach and measured its activity towards human GPR83 using NanoBiT-based ligand−receptor binding assay and NanoBiT-based β-arrestin recruitment assay. The results showed that FAM237B indeed displayed lower activity than its paralog FAM237A in both binding and activation assays; however, it could cause significant activation effect on human GPR83 at the nanomolar range (1‒10 nM). Thus, FAM237B might be another endogenous agonist for GPR83.