Molecular cloning and functional characterization of V2 [8-lysine] vasopressin and oxytocin receptors from a pig kidney cell line

Abstract

[Arg8]vasopressin and oxytocin are the two main members of the neurohypophysial hormone family found to be present in nearly all mammals. [Lys8]vasopressin ([Lys8]VP) has been identified as the antidiuretic hormone in pig and some marsupial families. The porcine-derived kidney epithelial cell line, LLC-PK1, expresses both [Lys8]VP receptors coupled to the activation of adenylate cyclase (V2 receptors) and oxytocin receptors. Here we report the molecular cloning of the V2 [Lys8]VP receptor and the oxytocin receptor from LLC-PK1 cells. The cloned V2 [Lys8]VP receptor differs from human and rat V2 [Arg8] receptors mainly in its N-terminal region, in residues located in the extracellular loops and in intracellular phosphorylation sites. When expressed in COS7 cells, the V2 [Lys8]VP receptor exhibits the relative order of ligand affinity [Lys8]VP = [Arg8]VP >> 1-deamino[D-Arg8]VP > or = oxytocin and adenylate-cyclase stimulation, expected for the porcine V2 [Lys8]VP receptor but different from V2 [Arg8]VP receptors. Adenylate-cyclase activation by [Lys8]VP was inhibited in COS7 cells by a V2 antagonist. The cloned oxytocin receptor exhibits in COS7 cells a ligand specificity typical of mammalian oxytocin receptors. mRNA-distribution analysis revealed a single 5.5-kb transcript in the uterus from pregnant guinea pig.