Mu-delta opioid receptor functional interaction: Insight using receptor-G protein fusions.
Fusion proteins between a receptor and a pertussis toxin-insensitive G(i)alpha subunit were used to gain insight into the molecular interactions that take place upon mu and delta opioid receptor heterodimerization. When mu opioid receptor-G(i1)alpha fusions were coexpressed with nonfused delta opioid receptors in human embryonic kidney 293 cells, or vice versa, receptor heterodimers were detected by coimmunoprecipitation. In pertussis toxin-treated cells, receptor coexpression decreased the amount of guanosine 5'-O-(3-[35S]thio)triphosphate ([35S]GTPgammaS) incorporated in the fused G alpha protein after the addition of agonists specific for the receptor-G(i1)alpha fusion. In addition, activation of the G alpha protein occurred in heterodimers upon addition of an agonist specific for the nonfused receptor. It remained unaffected by an inverse agonist specific for the receptor-G(i1)alpha fusion. These data suggest that signaling through the receptor-G(i1)alpha fusion protein is impaired in heterodimers and support a mechanism in which activation of the G alpha subunit is promoted by a direct interaction with the nonfused receptor. Alternatively, receptor coexpression did not modify the ligand binding properties for the high-affinity state of the receptor-G(i1)alpha fusion nor the EC50 values for agonist-induced [35S]GTPgammaS incorporation in the G(i1)alpha subunit. In addition, no binding competition was observed between delta and mu ligands. Together, the data point to mu-delta opioid receptor heterodimers formed by contact interactions between monomers that retain their structural integrity.
Snook
Laelie A
LA
Unité Mixte de Recherche 7104, Institut de Génétique et Biologie Moléculaire et Cellulaire, Centre National de la Recherche Scientifique/Institut National de la Santé et de la Recherche Médicale/Université Louis Pasteur, Illkirch, France.
Milligan
Graeme
G
Kieffer
Brigitte L
BL
Massotte
Dominique
D
D000701:Analgesics, Opioid / Q000494:pharmacology; D001549:Benzamides / Q000494:pharmacology; D001665:Binding Sites; D001667:Binding, Competitive / Q000187:drug effects; D015153:Blotting, Western; D002460:Cell Line; D002462:Cell Membrane / Q000502:physiology; D002478:Cells, Cultured; D020875:Enkephalin, Ala(2)-MePhe(4)-Gly(5)- / Q000494:pharmacology; D019204:GTP-Binding Proteins / Q000819:agonists / Q000502:physiology*; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000378:metabolism; D006801:Humans; D047468:Immunoprecipitation; D008024:Ligands; D009842:Oligopeptides / Q000494:pharmacology; D010455:Peptides / Q000494:pharmacology; D010879:Piperazines / Q000494:pharmacology; D017465:Receptors, Opioid, delta / Q000819:agonists / Q000502:physiology*; D017450:Receptors, Opioid, mu / Q000819:agonists / Q000502:physiology*; D011993:Recombinant Fusion Proteins / Q000494:pharmacology
D000701:Analgesics, Opioid; D001549:Benzamides; D008024:Ligands; D009842:Oligopeptides; D010455:Peptides; D010879:Piperazines; D017465:Receptors, Opioid, delta; D017450:Receptors, Opioid, mu; D011993:Recombinant Fusion Proteins; D020875:Enkephalin, Ala(2)-MePhe(4)-Gly(5)-; C060209:deltorphin II, Ala(2)-; C088464:4-(alpha-(4-allyl-2,5-dimethyl-1-piperazinyl)-3-methoxybenzyl)-N,N-diethylbenzamide; D016244:Guanosine 5'-O-(3-Thiotriphosphate); C009862:connective tissue-activating peptide; D019204:GTP-Binding Proteins