Clinical and Experimental Pharmacology and Physiology (2004) 31, 828–832 Annual Scientific Meeting of ASCEPT 2003 IDENTIFICATION AND CHARACTERIZATION OF THE MOUSE AND RAT RELAXIN RECEPTORS AS THE NOVEL ORTHOLOGUES OF HUMAN LEUCINE-RICH REPEAT-CONTAINING G-PROTEIN-COUPLED RECEPTOR 7 DJ Scott,* S Layfield,* A Riesewijk, † H Morita, ‡ GW Tregear* and RAD Bathgate* *Howard Florey Institute, University of Melbourne, Parkville, Victoria, Australia, † Department of Target Discovery, NV Organon, Oss, The Netherlands and ‡ Division of Reproductive Biology, Department of Gynecology and Obstetrics, Stanford University School of Medicine, Stanford, California, USA SUMMARY 1. Relaxin is an extracellular matrix (ECM)-remodelling hormone that is functionally important in reproductive tissues, brain, lung and heart. 2. Recently, the human relaxin receptor was identified as leucine-rich repeat-containing G-protein-coupled receptor 7 (LGR7). 3. Using human LGR7 as a template, we identified mouse and rat LGR7 orthologues in the Celera and National Centre for Biotechnology Information databases. 4. At the protein level, mouse and rat LGR7 share 85.2 and 85.7% identity with human LGR7, respectively. 5. Mouse LGR7 mRNA was detected in all tissues where relaxin binding is observed. 6. Mouse and rat LGR7 bound [ 33 P]-relaxin with high affinity and, upon relaxin treatment, both receptors stimulated cAMP production in transfected HEK 293T cells. 7. These results indicate that mouse and rat LGR7 are the relaxin receptors in these species. 8. The actions of relaxin in rodents are well characterized, providing an established platform for research into the molecular pharmacology of the highly conserved relaxin receptor. Key words: extracellular matrix, insulin-like peptide 3 (INSL3), insulin-like peptides, leucine-rich repeat-containing G-protein-coupled receptor 7 (LGR7), pregnancy, relaxin. INTRODUCTION Originally discovered and named for its ability to ‘relax’ the pubic ligaments of female guinea-pigs, 1 relaxin is primarily classified as a hormone of pregnancy. The principal role of relaxin, as a modulator of extracellular matrix (ECM) remodelling, is most evident during pregnancy, when it serves to reorganize the ECM of the uterus and cervix as well as lengthening the pubic ligaments to accommodate parturition. 2 However, studies in mouse and rat demonstrate that the roles of relaxin extend further than the boundaries of pregnancy. Relaxin-knockout mice are unable to feed their young owing to insufficient nipple development 3 and exhibit increased general collagen fibrosis in organs such as the heart and lung as they age. 4,5 Relaxin is a heterodimeric hormone, with its A and B chains linked together by two disulphide bonds. 6 Because relaxin exhibits an insulin-related structure, the relaxin receptor was predicted to be a single transmembrane tyrosine kinase receptor comparable to the insulin receptor. However, recently it was discovered that relaxin will activate a cAMP-dependent signalling pathway after binding to a novel human G-protein-coupled receptor (GPCR), leucine-rich repeat-containing GPCR 7 (LGR7). 7 The LGR family members, including the gonadotropin and thyrotropin receptors, are grouped together because they contain large extracellular ectodomains containing leucine-rich repeats (LRR). 8 Both the LGR7 and the closely related insulin-like peptide 3 (INSL3) receptor (LGR8) are unique LGR because they possess a single low-density lipoprotein receptor-like cysteine-rich motif (LDLR- domain) at their amino termini. 9 Similar LGR containing LDLR- domains have been identified in snail (Lymnaea stagnalis) 10 and fly (Drosophila melanogaster; accession no.: NM_170236), indi- cating this type of LGR is an ancient type of GPCR that has been conserved throughout evolution. To complement the numerous studies on relaxin in mouse and rat with investigations into the relaxin receptor, LGR7 must be identified and characterized as the relaxin receptor in these species. Using human LGR7 as a template, we identified novel LGR7 sequences from mouse and rat in the Celera (http:// www.celera.com) and National Centre for Biotechnology Correspondence: Dr R Bathgate, Howard Florey Institute, University of Melbourne, Victoria 3010, Australia. Email: r.bathgate@hfi.unimelb.edu.au Presented at the 37th Annual Scientific Meeting of the Australasian Society of Clinical and Experimental Pharmacologists and Toxicologists, Sydney, 29 November–3 December 2003. Received 12 February 2004; revision 23 July 2004; accepted 25 July 2004.