2ND MESSENGER CASCADE SPECIFICITY AND PHARMACOLOGICAL SELECTIVITY OF THE HUMAN P-2Y1-PURINOCEPTOR

1 The coding sequence of the P-2Y1-purinoceptor was cloned from a huma n genomic library. 2 The open reading frame encodes a protein of 373 a mino acids that is 83% identical to the previously cloned chick and tu rkey P-2Y1-purinoceptor and is greater than or equal to 95% homologous to the recently cloned rat, mouse, and bovine P-2Y1-purinoceptors 3 T he human P-2Y1-purinoceptor was stably expressed in 1321N1 human astro cytoma cells using a retroviral vector. Although the P-2Y1-purinocepto r agonist, 2MeSATP, had no effect on inositol phosphate accumulation i n 1321N1 cells infected with the control virus, this agonist markedly stimulated inositol phosphate accumulation in cells infected with the P-2Y1-purinoceptor virus. No effect of 2MeSATP on cyclic AMP accumulat ion was observed in P-2Y1-receptor-expressing 1321N1 cells. 4 The phar macological selectivity of 18 purinoceptor agonists was established fo r the expressed human P-2Y1-purinoceptor, 2MeSATP was more potent than ATP but less potent than 2MeSADP. ADP also was more potent than ATP. A similar maximal effect was observed with most agonists tested. Howev er, alpha,beta-MeATP had no effect and 3'-NH2-3'-deoxyATP and A(2)P(4) were partial agonists. The order of potency of agonists for activatio n of the turkey P-2Y1-purinoceptor, also stably expressed in 1321N1 ce lls, was identical to that observed for the human P-2Y1-purinoceptor. 5 C6 glioma cells express a P-2Y-purinoceptor that inhibits adenylyl c yclase but does not activate phospholipase C. Expression of the human P-2Y1-purinoceptor in C6 cells conferred 2MeSATP-stimulated inositol l ipid hydrolysis to these cells. The phospholipase C-activating human P -2Y1-purinoceptor could be delineated from the endogenous P-2Y-purinoc eptor of C6 glioma cells by use of the P-2-purinoceptor antagonist, PP ADS, which blocks the P-2Y1-purinoceptor but does not block the endoge nous P-2Y- purinoceptor of C6 cells. P-2-purinoceptor agonists also ex hibited differential selectivities for activation of these two P-2Y-pu rinoceptors.