Molecular cloning and functional expression of a VIP-specific receptor

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Abstract
Three receptors for VIP and pituitary adenylate cyclase-activating peptide (PACAP) have been cloned and characterized: PAC1, with high affinity for PACAP, and VPAC1and VPAC2with equally high affinity for VIP and PACAP. The existence of a VIP-specific receptor (VIPs) in guinea pig (GP) teniae coli smooth muscle was previously surmised on the basis of functional studies, and its existence was confirmed by cloning of a partial NH2-terminal sequence. Here we report the cloning of the full-length cDNAs of two receptors, a VPAC2receptor from GP gastric smooth muscle and VIPsfrom GP teniae coli smooth muscle. The cDNA sequence of the VIPsencodes a 437-amino acid protein ( Mr49,560) that possesses 87% similarity to VPAC2receptors in rat and mouse and differs from the VPAC2receptor in GP gastric smooth muscle by only two amino-acid residues, F40F41in lieu of L40L41. In COS-1 cells transfected with the GP teniae coli smooth muscle receptor, only VIP bound with high affinity (IC501.4 nM) and stimulated cAMP formation with high potency (EC501 nM). In contrast, in COS-1 cells transfected with the GP gastric smooth muscle receptor, both VIP and PACAP bound with equally high affinity (IC502.3 nM) and stimulated cAMP with equally high potency (EC501.5 nM). We conclude that the receptor cloned from GP teniae coli smooth muscle is a VIPsdistinct from VPAC1and VPAC2receptors. The ligand specificity in this species is determined by a pair of adjacent phenylalanine residues (L40L41) in the NH2-terminal ligand-binding domain.