A HUMAN GENE ENCODES A PUTATIVE G-PROTEIN-COUPLED RECEPTOR HIGHLY EXPRESSED IN THE CENTRAL-NERVOUS-SYSTEM

The mammalian bombesin (Bn)-like neuropeptide receptors gastrin-releas ing peptide receptor (GRP-R) and neuromedin B receptor (NMB-R) transdu ce a variety of physiological signals that regulate secretion, growth, muscle contraction, chemotaxis and neuromodulation. We have used reve rse transcription-polymerase chain reaction (PCR) to isolate a cDNA fr om human brain mRNA, GPCR/CNS, that encodes a putative G protein-coupl ed receptor (GPCR) based upon the presence of the paradigmatic seven h eptahelical transmembrane domains in its predicted amino acid sequence . Analysis of the deduced protein sequence of GPCR/CNS reveals this pu tative receptor to be 98% identical to the deduced amino acid sequence of a recently reported gene product and minimally identical (similar to 23%) to both murine GRP-R and human endothelin-B (ET-B) receptor. O ur deduced protein sequence differs at 12 positions, scattered through out the open reading frame, relative to the original sequence. A 3.7 k b GPCR/CNS mRNA species is expressed in vivo in a tissue-specific mann er, with highest levels detected in brain and spinal cord, lower level s found in testis, placenta and liver, but no detectable expression ob served in any other tissue. Analysis of GPCR/CNS genomic clones reveal s that the human gene contains one intron that is about 21 kb in lengt h that divides the coding region into two exons and maps to human chro mosome 7q31. No specific binding is observed with either a newly ident ified ligand (DTyr(6),beta Ala(11),Phe(13),Nle(14)]Bn-(6-14)) having h igh affinity for all Bn receptor subtypes or Bn after GPCR/CNS is stab ly expressed in fibroblasts. No elevation in inositol trisphosphate is observed after the application of micromolar levels of either DPhe(6) ,beta Ala(11),Phe(13),Nle(14)]Bn-(6-14) or Bn, a concentration of agon ist known to activate all four known Bn receptor subtypes. When GPCR/C NS is expressed in Xenopus oocytes, no activation of the calcium-depen dent chloride channel is detected despite the addition of micromolar l evels of Bn peptide agonists. We conclude that the natural ligand for this receptor is none of the known naturally occurring Bn-like peptide s and the true agonist for GPCR/CNS remains to be elucidated. (C) 1998 Elsevier Science B.V.