TY - JOUR T1 - Genetic and Functional Analysis of Human P2X5 Reveals a Distinct Pattern of Exon 10 Polymorphism with Predominant Expression of the Nonfunctional Receptor Isoform JF - Molecular Pharmacology JO - Mol Pharmacol SP - 953 LP - 960 DO - 10.1124/mol.110.063636 VL - 77 IS - 6 AU - Smita Kotnis AU - Brendan Bingham AU - Dmitry V. Vasilyev AU - Scott W. Miller AU - Yuchen Bai AU - Sarita Yeola AU - Pranab K. Chanda AU - Mark R. Bowlby AU - Edward J. Kaftan AU - Tarek A. Samad AU - Garth T. Whiteside Y1 - 2010/06/01 UR - http://molpharm.aspetjournals.org/content/77/6/953.abstract N2 - P2X5 is a member of the P2X family of ATP-gated nonselective cation channels, which exist as trimeric assemblies. P2X5 is believed to trimerize with another member of this family, P2X1. We investigated the single-nucleotide polymorphism (SNP) at the 3′ splice site of exon 10 of the human P2X5 gene. As reported previously, presence of a T at the SNP location results in inclusion of exon 10 in the mature transcript, whereas exon 10 is excluded when a G is present at this location. Our genotyping of human DNA samples reveals predominance of the G-bearing allele, which was exclusively present in DNA samples from white American, Middle Eastern, and Chinese donors. Samples from African American donors were polymorphic, with the G allele more frequent. Reverse transcription-polymerase chain reaction analysis of lymphocytes demonstrated a 100% positive correlation between genotype and P2X5 transcript. Immunostaining of P2X1/P2X5 stably coexpressing cell lines showed full-length P2X5 to be expressed at the cell surface and the exon 10-deleted isoform to be cytoplasmic. Fluorometric imaging-based pharmacological characterization indicated a ligand-dependent increase in intracellular calcium in 1321N1 astrocytoma cells transiently expressing full-length P2X5 but not the exon 10-deleted isoform. Likewise, electrophysiological analysis showed robust ATP-evoked currents when full-length but not the exon 10-deleted isoform of P2X5 was expressed. Taken together, our findings indicate that most humans express only a nonfunctional isoform of P2X5, which is in stark contrast to what is seen in other vertebrate species in which P2X5 has been studied, from which only the full-length isoform is known.Copyright © 2010 The American Society for Pharmacology and Experimental Therapeutics ER -