Expression of the third intracellular loop of the delta-opioid receptor inhibits signaling by opioid receptors and other G protein-coupled receptors.
To explore the feasibility of developing inhibitors of signaling by opioid receptors and other G protein-coupled receptors (GPCRs) that use the same G protein pool, we investigated the capacity of a minigene encoding the third intracellular loop of the delta-opioid receptor (delta-i3L) to act as competitive antagonist of the receptor-G protein interface interaction. In delta-i3L-expressing cells, the peptide blocked high-affinity agonist binding to both the delta- and the mu-opioid (delta-OR and mu-OR) and attenuated opioid and alpha2-adrenergic receptor (alpha2AR)-dependent [35S]guanosine-5'-O-(3-thio)triphosphate binding. Furthermore, delta-i3L expression resulted in inhibition of delta-, mu-OR-, and alpha2AR-receptor-mediated cAMP accumulation, whereas the cAMP response produced by activation of the beta2-adrenergic receptor was unaffected, suggesting that the inhibitory effects of delta-i3L expression were selective for Gi/Go proteins. Moreover, although delta-i3L expression also attenuated drastically phospholipase C accumulation and Ca2+ release following mu- and delta-OR stimulation, it failed to inhibit carbachol-mediated stimulation of inositol phosphate accumulation in M1-muscarinic receptor-expressing human embryonic kidney 293 cells. Finally, we also examined the effects of delta-i3L expression on the regulation of the extracellular signal-regulated kinase (ERK) mitogen-activated protein kinase pathway. Our results demonstrate that, although ERK activation by mu- and delta-ORs is attenuated by the presence of delta-i3L, ERK activation mediated by alpha2AR remained unaffected. Collectively, our data demonstrate that the delta-i3L can be used as potent inhibitor of G protein signaling for various GPCRs that use a common pool of G proteins.
Morou
Evangelia
E
Laboratory of Cellular Signaling and Molecular Pharmacology, Institute of Biology, National Center for Scientific Research "Demokritos", 15310 Ag. Paraskevi, Athens, Greece.
Georgoussi
Zafiroula
Z
D002118:Calcium / Q000032:analysis / Q000378:metabolism; D002460:Cell Line; D000242:Cyclic AMP / Q000037:antagonists & inhibitors / Q000096:biosynthesis; D019206:GTP-Binding Protein alpha Subunits, Gi-Go / Q000378:metabolism; D019204:GTP-Binding Proteins / Q000037:antagonists & inhibitors*; D006801:Humans; D008956:Models, Chemical; D009292:Narcotic Antagonists; D009842:Oligopeptides / Q000378:metabolism / Q000494:pharmacology; D010716:Phosphatidylinositols / Q000032:analysis / Q000378:metabolism; D017465:Receptors, Opioid, delta / Q000819:agonists / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects*
D009292:Narcotic Antagonists; D009842:Oligopeptides; D010716:Phosphatidylinositols; D017465:Receptors, Opioid, delta; D000242:Cyclic AMP; D019204:GTP-Binding Proteins; D019206:GTP-Binding Protein alpha Subunits, Gi-Go; D002118:Calcium