M. Thibonnier et al., MOLECULAR-CLONING, SEQUENCING, AND FUNCTIONAL EXPRESSION OF A CDNA-ENCODING THE HUMAN-V(1A) VASOPRESSIN RECEPTOR, The Journal of biological chemistry, 269(5), 1994, pp. 3304-3310
Vasopressin (AVP), the antidiuretic hormone, is a cyclic nonapeptide t
hat acts through binding to G protein-coupled specific membrane recept
ors pharmacologically divided into three subtypes (V1a, V1b, and V2) l
inked to distinct second messengers. Within the family of human AVP re
ceptors, the V2 AVP receptor has been cloned, but the structure of the
human V1a and V1b AVP receptors remains unknown. We report here the s
tructure and functional expression of a human V1a AVP receptor complem
entary DNA isolated from human liver cDNA libraries. Cloning and seque
ncing of a full-length clone isolated a 1472-nucleotide sequence encod
ing a 418-amino acid polypeptide with seven putative transmembrane dom
ains typical of G protein-coupled receptors. Amino acid sequence ident
ity with the rat liver V1a AVP receptor, the human and rat V2 AVP rece
ptors, and the human oxytocin receptor was 72, 36, 37, and 45%, respec
tively. Functional characterization of the cloned receptor was done by
transient expression in COS-7 cells and stable expression in Chinese
hamster ovary cells. Localization of the expressed receptor at the cel
lular surface was illustrated by using the fluorescent linear analog p
henylacetyl-D-Tyr(Et)-Phe-Gln-Asn-Lys-Pro-Arg-NH2 coupled to fluoresce
in-avidin by dodecabiotin. Competition binding experiments with cetyl-
D-Tyr(Et)-Phe-Val-Asn-Lys-Pro-[I-125]Tyr-NH2 and AVP analogs revealed
high affinity specific binding sites of the V1a subtype. Saturation bi
nding experiments with [H-3]AVP confirmed the presence of a single cla
ss of high affinity binding sites. Measurement of AVP-induced inositol
phosphate production and calcium mobilization confirmed that the expr
essed V1a AVP receptor is coupled to phospholipase C via a pertussis t
oxin-insensitive pathway. Thus, the human V1a AVP receptor belongs to'
'the superfamily of seven-transmembrane segment receptors with a signi
ficant sequence identity with the other members of the AVP-oxytocin fa
mily of receptors.