TY - JOUR T1 - Distinct Signaling Patterns of Allosteric Antagonism at the P2Y<sub>1</sub> Receptor JF - Molecular Pharmacology JO - Mol Pharmacol DO - 10.1124/mol.117.109660 SP - mol.117.109660 AU - Zhan-Guo Gao AU - Kenneth A. Jacobson Y1 - 2017/01/01 UR - http://molpharm.aspetjournals.org/content/early/2017/09/01/mol.117.109660.abstract N2 - Traditionally, GPCR antagonists are classified as competitive or non-competitive and surmountable or insurmountable based on functional antagonism. P2Y1 receptor (P2Y1R) structures showed two antagonists binding to two spatially distinct sites: nucleotide MRS2500 (orthosteric, contacting the helical bundle) and urea BPTU (allosteric, on the external receptor surface). However, the nature of their P2Y1R antagonism has not been characterized. Here we characterized BPTU antagonism at various signaling pathways activated by structurally diverse agonists. BPTU rightward-shifted the concentration-response curves of both 2MeSADP and MRS2365 (5'-diphosphates) in some signaling events, such as ERK1/2 and label-free), in a parallel manner without affecting the maximum agonist effect (Emax), but antagonized insurmountably (suppressed agonist Emax) in signaling events such as GTPγS binding and β-arrestin2 recruitment. However, with dinucleotide Ap4A as an agonist, BPTU suppressed the Emax insurmountably in all signaling pathways. By comparison, MRS2500 behaved as a surmountable antagonist rightward-shifting concentration-response curves of all three agonists in a parallel manner for all signaling pathways measured. Thus, we demonstrated a previously undocumented phenomenon that P2Y1R antagonism patterns could vary in different signaling pathways, which could be related to conformational selection, signaling amplification and probe dependence. This phenomenon may apply generally to other receptors considering that antagonism by a specific ligand is often not compared at multiple signaling pathways. Thus, antagonism can be surmountable or insurmountable depending signaling pathways measured and agonists used, which should be of broad relevance to drug discovery and disease treatment. ER -