AN EXTRACELLULAR CONGENITAL NEPHROGENIC DIABETES-INSIPIDUS MUTATION OF THE VASOPRESSIN RECEPTOR REDUCES CELL-SURFACE EXPRESSION, AFFINITY FOR LIGAND, AND COUPLING TO THE G(S)/ADENYLYL CYCLASE SYSTEM

The mutation of the type-2 vasopressin receptor (V2R) apparently responsible for X-linked congenital nephrogenic diabetes insipidus (CNDI) in the Q3 family consists of a T to C transition in codon 113, causing the change of Arg-113 to Trp. Arg-113 is located in the putative first extracellular loop of the V2R next to a frequently conserved Cys thought to interact via a disulfide bridge with a Cys of the second extracellular loop. The present study explored whether this mutation may account for the CNDI phenotype. The mutation was excised from the genomic DNA of a Q3 patient and introduced into the V2R cDNA, which was then placed into an expression plasmid and transfected into COS cells for transient expression and murine L cells for stable expression. Studies with L cells expressing similar levels of wild type and Q3 receptors showed that the mutant receptor has a 20-fold reduced affinity for arginine vasopressin (AVP) and stimulates adenylyl cyclase with an EC(50) that is increased by a factor of about 60-fold. The same shift in the EC(50) for adenylyl cyclase stimulation was obtained when deamino[8-D-Arg]vasopressin was substituted for AVP. Studies with COS cells revealed that at equal levels of transfected DNA, the mutant receptor is expressed at lower levels (about 20%) than the wild type receptor, indicating that the mutation hinders the transport of the receptor to the cell membrane. Studies with L cells showed that a reduction of wild type receptor may lead to a right shift in the EC(50) for AVP stimulation of adenylyl cyclase and a reduction of the extent to which adenylyl cyclase is stimulated. We conclude that the combination of lowered affinity for ligand and diminished ability to stimulate adenylyl cyclase and to reach the cell surface may well account for the CNDI syndrome of Q3 patients, i.e. for their unresponsiveness to the antidiuretic action of AVP and to clinically administered V2R selective analog deamino[8-D-Arg]vasopressin.