title: INSL5 is a high affinity specific agonist for GPCR142 (GPR100).
abstract: Insulin-like peptide 5 (INSL5) is a peptide that belongs to the relaxin/insulin family, and its receptor has not been identified. In this report, we demonstrate that INSL5 is a specific agonist for GPCR142. Human INSL5 displaces the binding of (125)I-relaxin-3 to GPCR142 with a high affinity (K(i) = 1.5 nM). In a saturation binding assay, (125)I-INSL5 binds GPCR142 with a K(d) value of 2.5 nM. In functional guanosine (gamma-thio)-triphosphate binding and cAMP accumulation assays, INSL5 potently activates GPCR142 with EC(50) values of 1.3 and 1.2 nM, respectively. In addition, INSL5 stimulates Ca(2+) mobilization in HEK293 cells expressing GPCR142 and G alpha(16). Overall, INSL5 behaves as an agonist for GPCR142 similar to relaxin-3. However, unlike relaxin-3, which is also a potent agonist for GPCR135 and LGR7, INSL5 does not activate either GPCR135 or LGR7. INSL5 inhibits (125)I-relaxin-3 binding to GPCR135 with a low potency (K(i) = 500 nM). A functional assay shows that INSL5 (1 microm) is a weak antagonist for GPCR135. In addition, INSL5 (up to 1 microm) shows no affinity or activity at LGR7 or LGR8 either in a binding assay or a bio-functional assay. Previously, we have demonstrated that GPCR142 mRNA is expressed in peripheral tissues, particularly in the colon. Here we show that INSL5 mRNA is expressed in many peripheral tissues, similar to GPCR142. The high affinity interaction between INSL5 and GPCR142 coupled with their co-evolution and partially overlapping tissue expression patterns strongly suggest that INSL5 is an endogenous ligand for GPCR142.
authors.affiliation: Johnson & Johnson Pharmaceutical Research and Development, San Diego, California 92121, USA. cliu9@prdus.jnj.com
authors.forename: Changlu
authors.initials: C
authors.lastname: Liu
authors.forename: Chester
authors.initials: C
authors.lastname: Kuei
authors.forename: Steven
authors.initials: S
authors.lastname: Sutton
authors.forename: Jingcai
authors.initials: J
authors.lastname: Chen
authors.forename: Pascal
authors.initials: P
authors.lastname: Bonaventure
authors.forename: Jiejun
authors.initials: J
authors.lastname: Wu
authors.forename: Diane
authors.initials: D
authors.lastname: Nepomuceno
authors.forename: Fredrik
authors.initials: F
authors.lastname: Kamme
authors.forename: Da-Thao
authors.initials: DT
authors.lastname: Tran
authors.forename: Jessica
authors.initials: J
authors.lastname: Zhu
authors.forename: Tracey
authors.initials: T
authors.lastname: Wilkinson
authors.forename: Ross
authors.initials: R
authors.lastname: Bathgate
authors.forename: Elo
authors.initials: E
authors.lastname: Eriste
authors.forename: Rannar
authors.initials: R
authors.lastname: Sillard
authors.forename: Timothy W
authors.initials: TW
authors.lastname: Lovenberg
mesh_terms: D000595:Amino Acid Sequence; D006801:Humans; D007328:Insulin / Q000378:metabolism*; D008024:Ligands; D008969:Molecular Sequence Data; D009928:Organ Specificity; D011485:Protein Binding; D011506:Proteins / Q000378:metabolism*; D012333:RNA, Messenger / Q000096:biosynthesis; D011869:Radioligand Assay; D043562:Receptors, G-Protein-Coupled; D018000:Receptors, Peptide / Q000819:agonists / Q000378:metabolism*; D016415:Sequence Alignment
chemical_list: D007328:Insulin; C084797:Leydig insulin-like protein; D008024:Ligands; D011506:Proteins; D012333:RNA, Messenger; C480239:RXFP4 protein, human; D043562:Receptors, G-Protein-Coupled; D018000:Receptors, Peptide