title: P2X1 purinoceptor in human platelets. Molecular cloning and functional characterization after heterologous expression.
abstract: ADP is an important physiological platelet agonist. The molecular identity of the ADP receptor(s) in human platelets, however, is still unclear. Although P2T purinoceptor was believed to be the ligand-gated cation channel for ADP in human platelets, recent patch clamp studies now suggest it is P2X1 type. In the present study, we have cloned a cDNA encoding a P2X1 purinoceptor from human platelets using degenerate reverse transcription and polymerase chain reaction. Northern blotting with a P2X1-specific probe revealed a band of 1.8 kilobases in human platelets as well as in several megakaryoblastic cell lines. 1321N1 human astrocytoma cells expressing the cloned P2X1 cDNA exhibited both ATP- and ADP-stimulated Ca2+ influx that could be blocked by the purinoceptor antagonist pyridoxalphosphate-6-azophenyl-2',4'-disulfonic acid and suramin. Additionally, a polyclonal antibody raised against glutathione-S-transferase-P2X1 fusion peptide reacted with a 70-kDa band on Western blot of human platelets. It is therefore concluded that functional P2X1 purinoceptors are present in human platelets.
authors.affiliation: Department of Thrombosis and Vascular Biology, Maryland Research Laboratories, Otsuka America Pharmaceutical, Inc., Rockville, Maryland 20850, USA. bings@mrl.oapi.com
authors.forename: B
authors.initials: B
authors.lastname: Sun
authors.forename: J
authors.initials: J
authors.lastname: Li
authors.forename: K
authors.initials: K
authors.lastname: Okahara
authors.forename: J
authors.initials: J
authors.lastname: Kambayashi
mesh_terms: D000244:Adenosine Diphosphate / Q000502:physiology; D000255:Adenosine Triphosphate / Q000502:physiology; D001254:Astrocytoma; D001483:Base Sequence; D001792:Blood Platelets / Q000502:physiology*; D002118:Calcium / Q000502:physiology; D003001:Cloning, Molecular; D003593:Cytoplasm / Q000378:metabolism; D006801:Humans; D015640:Ion Channel Gating; D008969:Molecular Sequence Data; D008970:Molecular Weight; D018048:Receptors, Purinergic P2 / Q000235:genetics* / Q000502:physiology; D058469:Receptors, Purinergic P2X; D011994:Recombinant Proteins; D014162:Transfection
chemical_list: D018048:Receptors, Purinergic P2; D058469:Receptors, Purinergic P2X; D011994:Recombinant Proteins; D000244:Adenosine Diphosphate; D000255:Adenosine Triphosphate; D002118:Calcium