Molecular cloning and characterization of two novel retinoic acid-inducible orphan G-protein-coupled receptors (GPRC5B and GPRC5C)

Using homology searching of public databases with a metabotropic glutamate receptor sequence from Caenorhabditis elegans, two novel protein sequences (named RAIG-2 (HGMW-approved symbol GPRC5B) and RAIG-3 (HGMW-approved symbo l GPRC5C) were identified containing seven putative transmembrane domains c haracteristic of G-protein-coupled receptors (GPCRs). RAIG-2 and RAIG-3 enc ode open reading frames of 403 and 442 amino acid polypeptides, respectivel y, and show 58% similarity to the recently identified retinoic acid inducib le gene-1 (RAIG-1, HGMW-approved symbol RAI3). Analysis of the three protei n sequences places them within the type 3 GPCR family; which includes metab otropic glutamate receptors, GABA, receptors, calcium-sensing receptors, an d pheromone receptors. However, in contrast to other type 3 GPCRs, RAIG-1, RAIG-2, and RAIG-3 have only short N-terminal domains. RAIG-2 and RAIG-3 cD NA sequences were cloned into the mammalian expression vector pcDNA3 with c -myc or HA epitope tags inserted at their N-termini, respectively. Transien t transfection experiments in HEK239T cells using these constructs demonstr ated RAIG-2 and RAIG-3 expression at the cell surface. Distribution profile s of mRNA expression obtained by semiquantitative TaqMan PCR analysis showe d RAIG-2 to be predominantly expressed in human brain areas and RAIG 3 to b e predominantly expressed in peripheral tissues. Tn addition, expression of RAIG-2 and RAIG-3 mRNA was increased following treatment with all-trans-re tinoic acid in a manner similar to Chat previously described for RAIG-1. Fi nally RAIG-2 was mapped to chromosome 16p12 (D16S405-D16S3045) and RAIG-3 t o chromosome 17q25 CD17S1352-D17S785). These results suggest that RAIG-1, R AIG-2, and RAIG-3 represent a novel family of retinoic acid-inducible recep tors, most closely related to the type 3 GPCR subfamily, and provide furthe r evidence for a linkage between retinoic acid and G-protein-coupled recept or signal transduction pathways. (C) 2000 Academic Press.