Elsevier

Neuropharmacology

Volume 50, Issue 4, March 2006, Pages 512-520
Neuropharmacology

Characterisation and differential expression of two very closely related G-protein-coupled receptors, GPR139 and GPR142, in mouse tissue and during mouse development

https://doi.org/10.1016/j.neuropharm.2005.11.003Get rights and content

Abstract

By searching the human and mouse genomic databases we found two G-protein-coupled receptors, GPR139 and GPR142, with characteristic motifs of the rhodopsin family of receptors. The gene for GPR139 maps to chromosome 7F1 of mouse and 16p12.3 of human and that for GPR142 to 11E2 of mouse and 17q25.1 of human. We isolated GPR139 from a cDNA library of adult mouse brain and GPR142 from a cDNA library of brains from 15-day-old mouse embryos. GPR139 mRNA was predominantly expressed in specific areas of human and mouse brains, whereas GPR142 mRNA showed a more ubiquitous expression both in the brain and in various peripheral glands and organs. A 50% identity and a 67% homology at the amino-acid level between the two receptors and only 20–25% identity with other G-protein-coupled receptors established them as a new subbranch within the phylogenetic tree and hints at a common or similar ligand(s). Preliminary results suggest that the cognate ligand is present in brain extracts and is, most likely, a small peptide. GPR139 signal transduction in Chinese hamster ovary cells requires coupling to an inhibitory G-protein and is mediated by phospholipase C. Dimer formation may be necessary for proper function.

Introduction

G-protein-coupled receptors (GPCRs) are the largest family of cell-surface receptors transducing signals from the extracellular environment to intracellular effectors. Ligands are of diverse molecular structure and size and include light, ions, amino acids, nucleotides, metabolic intermediates, lipids, peptides and proteins (Civelli, 2005). GPCRs are characterised by seven transmembrane helices separated by intra- and extracellular loops of variable lengths. Together with the extracellular amino terminus and the cytosolic carboxy terminus the loops determine ligand interactions and signal-transduction pathways (Bockaert and Pin, 1999). The classification of GPCRs into subfamilies is based on their homology within the heptahelical structure and partially on their interaction with ligands (Frederiksson et al., 2003a). GPCRs play key physiological roles, both during development and in the adult, and their dysfunction is implicated in several diseases (Hökfelt et al., 2003). This is reflected by the fact that about half of the current drugs, and certainly more in the future, are targeted to GPCRs. Discovery of novel receptors and their ligands is, therefore, of great interest for pharmacology.

In this paper we describe the identification, cloning and tissue distribution of two mouse GPCRs, GPR139 and GPR142, which belong to the rhodopsin family of GPCRs. They are highly related to each other, but differ strongly in expression patterns within the brain and in other tissues. A ligand present in brain extracts activated Ca2+ mobilisation in GPR139-expressing cells. Signalling was mediated by an inhibitory G-protein and by phospholipase C and required dimer formation for proper function.

Section snippets

Identification and cloning of mouse GPR139 and GPR142

By searching the human genomic database with known peptide GPCRs using TBLASTN we discovered GPR139 and GPR142. The conserved regions of the human receptors subsequently served to find the respective mouse orthologues. A full-length cDNA for mouse GPR139 was obtained by 5′- and 3′-rapid amplification of cDNA ends (RACE) from a RACE-ready cDNA of mouse brain (Ambion, Huntingdon, UK). Nested RACE primers were designed over an internal XbaI site at position 300 of the mouse nucleotide sequence (AY485344

Identification, cloning and structure of mouse GPR139 and GPR142

The human genomic database was searched for novel sequences related to peptide-binding GPCRs using TBLASTN. Two related sequences were found in the human genome, one located on chromosome 16p12.3 and the other on 17q25.1. They shared 50% amino-acid identity in a region behind a splice site within the first transmembrane region. With their help mouse orthologues were also discovered, located on chromosomes 7F1 and 11E2. While this work was in progress human and mouse sequences were predicted and

Discussion

The mouse G-protein-coupled receptors GPR139 and GPR142 show with 94% and 74% identity at the amino-acid level, respectively, a high degree of relatedness to their human counterparts, which suggests a conserved function in evolution. The two receptors in mouse share 50% of their amino acids with each other, suggestive of a common ligand. The relatedness of 20–25% to other members of the rhodopsin family included receptors for thyrotrophin-releasing hormone, opioid peptides, somatostatin,

Acknowledgements

We thank J. Stables for kindly providing vectors containing apoaequorin and Gα16, A. Methner for the luciferase-reporter-gene constructs, A. Ignatov and M. Rezgaoui for critical comments on the manuscript and S. Hempel for help with the figures.

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