Human P2Y(2) receptor polymorphism: identification and pharmacological characterization of two allelic variants

1 In the process of cloning the human P2Y(2) receptor in order to establish 1321N1 cell lines expressing this receptor, we detected a gene polymorphis m characterized by an arginine 334 to cysteine 334 transition. 2 The frequency distribution of the polymorphism was studied in a European population. We observed that 66% of the tested persons are homozygotes R/R, 29% are heterozygotes R/C and 5% are homozygotes C/C. The frequency of the R allele was 0.8 versus 0.2 for the C allele. 3 We stably expressed each form of the human P2Y(2) receptor into 1321N1 ce lls and isolated clones by limiting dilution. The effects of nucleotides an d antagonists on inositol trisphosphate accumulation and cyclic AMP formati on were compared between the two cell lines. 4 The time-courses of inositol trisphosphate accumulation as well as concen tration-response curves characterizing the effects of UTP, ATP, AP(4)A and ATP gamma S were mostly similar, except for slight kinetic differences (slo wer time-course with the C-334 form). 5 The sensitivity to pertussis toxin of inositol trisphosphates accumulatio n was critically dependent on the agonist concentration and stimulation dur ation, suggesting the involvement of a G(i,0), protein during the early sti mulation by low nucleotide concentrations. No inhibition of cyclic AMP accu mulation could be detected. These properties were observed with both polymo rphic receptors.