title: Selective interactions of spinophilin with the C-terminal domains of the δ- and μ-opioid receptors and G proteins differentially modulate opioid receptor signaling.
abstract: Previous studies have shown that the intracellular domains of opioid receptors serve as platforms for the formation of a multi-component signaling complex consisting of various interacting partners (Leontiadis et al., 2009, Cell Signal. 21, 1218-1228; Georganta et al., 2010, Neuropharmacology, 59(3), 139-148). In the present study we demonstrate that spinophilin a dendritic-spine enriched scaffold protein associates with δ- and μ-opioid receptors (δ-ΟR, μ-OR) constitutively in HEK293 an interaction that is altered upon agonist administration and enhanced upon forskolin treatment for both μ-OR and δ-ΟR. Spinophilin association with the opioid receptors is mediated via the third intracellular loop and a conserved region of the C-terminal tails. The portion of spinophilin responsible for interaction with the δ-OR and μ-OR is narrowed to a region encompassing amino acids 151-444. Spinophilin, RGS4, Gα and Gβγ subunits of G proteins form a multi-protein complex using specific regions of spinophilin and a conserved amino acid stretch of the C-terminal tails of both δ-μ-ORs. Expression of spinophilin in HEK293 cells potentiated DPDPE-mediated adenylyl-cyclase inhibition of δ-OR leaving unaffected the levels of cAMP accumulation mediated by the μ-OR. Moreover, measurements of extracellular signal regulated kinase (ERK1,2) phosphorylation indicated that the presence of spinophilin attenuated agonist-driven ERK1,2 phosphorylation mediated upon activation of the δ-OR but not the μ-OR. Collectively, these findings suggest that spinophilin associates with both δ- and μ-ΟR and G protein subunits in HEK293 cells participating in a multimeric signaling complex that displays a differential regulatory role in opioid receptor signaling.
authors.affiliation: Laboratory of Cellular Signalling and Molecular Pharmacology, Institute of Biosciences and Applications, National Centre for Scientific Research Demokritos, Athens, Greece.
authors.forename: Danai-Dionysia
authors.initials: DD
authors.lastname: Fourla
authors.forename: Maria-Pagona
authors.initials: MP
authors.lastname: Papakonstantinou
authors.forename: Stavroula-Maria
authors.initials: SM
authors.lastname: Vrana
authors.forename: Zafiroula
authors.initials: Z
authors.lastname: Georgoussi
mesh_terms: D000262:Adenylyl Cyclases / Q000378:metabolism; D005576:Colforsin / Q000494:pharmacology; D000242:Cyclic AMP / Q000378:metabolism; D020881:Enkephalin, D-Penicillamine (2,5)- / Q000494:pharmacology; D019204:GTP-Binding Proteins / Q000378:metabolism*; D057809:HEK293 Cells; D006801:Humans; D008840:Microfilament Proteins / Q000235:genetics / Q000378:metabolism*; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism; D048052:Mitogen-Activated Protein Kinase 3 / Q000378:metabolism; D009419:Nerve Tissue Proteins / Q000235:genetics / Q000378:metabolism*; D010766:Phosphorylation / Q000187:drug effects; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D021122:Protein Subunits / Q000378:metabolism; D017465:Receptors, Opioid, delta / Q000819:agonists / Q000378:metabolism*; D017450:Receptors, Opioid, mu / Q000819:agonists / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects; D000099273:Neurabins
chemical_list: D000262:Adenylyl Cyclases; D005576:Colforsin; D000242:Cyclic AMP; D020881:Enkephalin, D-Penicillamine (2,5)-; D019204:GTP-Binding Proteins; D008840:Microfilament Proteins; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D009419:Nerve Tissue Proteins; D021122:Protein Subunits; D017465:Receptors, Opioid, delta; D017450:Receptors, Opioid, mu; D000099273:Neurabins