MOLECULAR-CLONING AND FUNCTIONAL-ANALYSIS OF A NOVEL P-2 NUCLEOTIDE RECEPTOR

The cDNA encoding a novel P-2 receptor was isolated from rat aortic smooth muscle cell library and functionally characterized. The cloned P-2 receptor exhibits structural features characteristic of the G protein-coupled receptor family and shows 44 and 38% amino acid identity with previously cloned rat P-2U and chicken P-2Y receptors, respectively. The cloned P-2 receptor is functionally coupled to phospholipase C but not to adenylate cyclase in C6 rat glioma cells transfected with the cloned P-2 expression vector. The rank order of agonist potency as judged by intracellular Ca2+ mobilization responses is UTP > ADP = 2-methylthioATP > ADP beta S > ATP = ATP gamma S, which is not compatible with any of the previously characterized P-2 receptor subtypes. The nonselective P-2 antagonists, suramin and reactive blue-2, inhibit nucleotide-induced phospholipase C activation in cells expressing the cloned P-2 receptor. The cloned P-2 receptor mRNA is abundantly expressed in various rat tissues including lung, stomach, intestine, spleen, mesentery, heart, and, most prominently, aorta. The results indicate that the novel metabotropic P-2 receptor has pharmacological characteristics distinct from any of P-2 receptor subtypes thus far identified and suggest the existence of a novel regulatory system by extracellular nucleotides of potential significance.