P 2 Y 13 : Identification and Characterization of a Novel G i-Coupled ADP Receptor from Human and Mouse

We have identified an orphan G protein-coupled receptor, SP174, that shares a high degree of homology with the recently described ADP receptor P2Y12. mRNA for SP174 is abundant in the brain and in cells of the immune system. In the present study, we demonstrate that SP174 is also a receptor for ADP, which is coupled to G i. ADP potently stimulates SP174 with an EC50 of 60 nM, and other related nucleotides are active as well, with a rank order of potency 2-methylthio-ADP tetrasodium adenosine 5 -O-2-(thio)diphosphate 2-methylthio-ATP tetrasodium ADP AP3A ATP IDP. This pharmacological profile is similar to that for P2Y12. We have also identified the murine homolog of SP174, which exhibits 75% homology to the human receptor. ADP is also a potent agonist at the murine receptor, and its pharmacological profile is similar to its human counterpart, but ADP and related nucleotides are more potent at the murine receptor than the human receptor. In keeping with the general nomenclature for the purinergic receptors, we propose designating this novel receptor P2Y13. P2Y receptors are G protein-coupled receptors that respond to the presence of extracellular nucleotides. Both purine and pyrimidine nucleotides can modulate a variety of physiological functions by interaction with P2Y receptors (Harden et al., 1995; Burnstock, 1997; Ralevic and Burnstock, 1998). Six mammalian P2Y receptors have been cloned so far, including P2Y1, P2Y2, P2Y4, P2Y6, P2Y11, and more recently, P2Y12 (Burnstock, 1997; Communi et al., 1997; Ralevic and Burnstock, 1998; Hollopeter et al., 2001; Zhang et al., 2001). P2Y1, P2Y2, and P2Y6 couple to the activation of phospholipase C (PLC); P2Y11 couples to the activation of both PLC and the adenylyl cyclase pathways, whereas human P2Y4 couples to adenylyl cyclase pathways at the early stage and PLC at a later stage (Communi et al., 1996; Ralevic and Burnstock, 1998). P2Y1 is selectively activated by ADP with ATP, being either a partial agonist or an antagonist; P2Y2 is activated equipotently by ATP and UTP; human P2Y11 is selectively activated by ATP; human P2Y6 is selectively activated by UDP, whereas rat P2Y6 is selectively activated by UTP; and human P2Y4 is activated selectively by UTP, whereas rat and murine P2Y4 are activated equipotently by ATP and UTP (Harden et al., 1995; Burnstock, 1997; Ralevic and Burnstock, 1998). In contrast, the P2Y12 receptor recently described by us (Zhang et al., 2001) and Hollopeter et al. (2001) is potently activated by ADP and is coupled to the inhibition of adenylyl cyclase activity through the G i class of G proteins. Analysis of the expression profile of P2Y12 receptor mRNA revealed that it is expressed at high levels in platelets, in addition to brain tissue. The data presented by Hollopeter et al. (2001) and the analysis of platelet function in P2Y12 null mice by Foster et al. (2001) clearly indicate that P2Y12 represents the long sought-after platelet ADP receptor. Furthermore, these studies reveal that P2Y12 is the molecular target of the important antithrombotic drug clopidogrel (Boyer et al., 1993; Daniel et al., 1998; Foster et al., 2001; Hollopeter et