scholarly journals Conformational specificity of opioid receptors is determined by subcellular location irrespective of agonist

eLife ◽  
2021 ◽  
Vol 10 ◽  
Author(s):  
Stephanie E Crilly ◽  
Wooree Ko ◽  
Zara Y Weinberg ◽  
Manojkumar A Puthenveedu
Keyword(s):  
G Protein ◽  
Opioid Receptor ◽  
Opioid Receptors ◽  
Neuronal Cells ◽  
Local Environment ◽  
Subcellular Location ◽  
Neuroendocrine Cells ◽  
Spatially Resolved ◽  
Δ Opioid Receptor ◽  
G Protein Coupled

The prevailing model for the variety in drug responses is that they stabilize distinct active states of their G protein-coupled receptor (GPCR) targets, allowing coupling to different effectors. However, whether the same ligand generates different GPCR active states based on the immediate environment of receptors is not known. Here we address this question using spatially resolved imaging of conformational biosensors that read out distinct active conformations of the δ-opioid receptor (DOR), a physiologically relevant GPCR localized to Golgi and the surface in neuronal cells. We show that Golgi and surface pools of DOR both inhibit cAMP, but engage distinct conformational biosensors in response to the same ligand in rat neuroendocrine cells. Further, DOR recruits arrestins on the surface but not the Golgi. Our results suggest that the local environment determines the active states of receptors for any given drug, allowing GPCRs to couple to different effectors at different subcellular locations.

scholarly journals Conformational specificity of opioid receptors is determined by subcellular location irrespective of agonist

2021 ◽  
Author(s):  
Stephanie E. Crilly ◽  
Wooree Ko ◽  
Zara Y. Weinberg ◽  
Manojkumar A. Puthenveedu
Keyword(s):  
Plasma Membrane ◽  
G Protein ◽  
Opioid Receptors ◽  
Live Cell Imaging ◽  
Cell Imaging ◽  
Subcellular Location ◽  
Unanswered Question ◽  
Receptor Coupling ◽  
Δ Opioid Receptor ◽  
G Protein Coupled

AbstractThe prevailing model for the variety in drug responses is that they stabilize distinct active states of their G protein-coupled receptor (GPCR) targets, allowing coupling to different effectors. However, whether the same ligand can produce different GPCR active states based on the environment of receptors in cells is a fundamental unanswered question. Here we address this question using live cell imaging of conformational biosensors that read out distinct active conformations of the δ-opioid receptor (DOR), a physiologically relevant GPCR localized to Golgi and the surface in neurons. We show that, although Golgi and surface pools of DOR regulated cAMP, the two pools engaged distinct conformational biosensors in response to the same ligand. Further, DOR recruited arrestin on the plasma membrane but not the Golgi. Our results suggest that the same agonist drives different conformations of a GPCR at different locations, allowing receptor coupling to distinct effectors at different locations.

scholarly journals Agonist activation of p42 and p44 mitogen-activated protein kinases following expression of the mouse δ opioid receptor in Rat-1 fibroblasts: effects of receptor expression levels and comparisons with G-protein activation

1996 ◽  
Vol 320 (1) ◽  
pp. 227-235 ◽  
Author(s):  
Andrew R BURT ◽  
I. Craig CARR ◽  
Ian MULLANEY ◽  
Neil G. ANDERSON ◽  
Graeme MILLIGAN
Keyword(s):  
Membrane Protein ◽  
G Protein ◽  
Opioid Receptor ◽  
Gtpase Activity ◽  
Protein Activation ◽  
G Protein Activation ◽  
Mitogen Activated Protein ◽  
Δ Opioid Receptor ◽  
G Protein Coupled ◽  
Stimulation Of

Rat-1 fibroblasts were transfected with a cDNA encoding the mouse Δ opioid receptor. Two separate clones, D2 (which expressed some 6 pmol of the receptor/mg of membrane protein) and DOE (which expressed some 0.2 pmol/mg of membrane protein), were examined in detail. With membranes from both clones, the opioid agonist [D-Ala2]leucine enkephalin (DADLE) caused stimulation of high-affinity GTPase activity and of the binding of guanosine 5´-[γ-[35S]thio]triphosphate, and inhibition of forskolin-amplified adenylate cyclase activity. DADLE also induced phosphorylation and activation of both the p42MAPK (42 kDa isoform) and p44MAPK (44 kDa isoform) members of the mitogen-activated protein kinase (MAP kinase) family. All of these effects of DADLE were prevented in both clones by pretreatment of the cells with pertussis toxin. The maximal response that could be produced by DADLE in direct assays of G-protein activation were substantially greater in clone D2 than in clone DOE, but in both clones essentially full phosphorylation of both p42MAPK and p44MAPK could be achieved. EC50 values for DADLE stimulation of GTPase activity and for activation of p44MAPK were substantially lower in clone D2 than in clone DOE. Moreover, in both clones the EC50 value for DADLE stimulation of p44MAPK was substantially lower than that for stimulation of GTPase activity, and the Hill coefficients for agonist activation of p44MAPK (h > 1) displayed marked co-operativity whereas those for G-protein activation did not (h 0.8–1.0). DADLE activation of p44MAPK showed more sustained kinetics in clone D2 than in clone DOE. By contrast, lysophosphatidic acid, acting at an endogenously expressed G-protein-coupled receptor, also activated p44MAPK in both clones in a pertussis toxin-sensitive manner, but both the kinetics and the concentration–response curve for activation of p44MAPK by this ligand were similar. As with other systems, maintained cellular levels of a cAMP analogue prevented the effects of both G-protein-coupled receptors on activation of p44MAPK. These results demonstrate for the first time that an opioid receptor, at least when expressed in Rat-1 fibroblasts, is able to initiate activation of the MAP kinase cascade in a Gi-dependent manner, and show that only a very small proportion of the cellular Gi population is required to be activated to result in full phosphorylation of the p42MAPK and p44MAPK MAP kinases.

scholarly journals Biased Opioid Ligands

Molecules ◽  
2020 ◽  
Vol 25 (18) ◽  
pp. 4257 ◽  
Author(s):  
Abdelfattah Faouzi ◽  
Balazs R. Varga ◽  
Susruta Majumdar
Keyword(s):  
G Protein ◽  
Opioid Receptor ◽  
Opioid Receptors ◽  
Signal Transduction Pathway ◽  
Opioid Peptide ◽  
Nop Receptor ◽  
Biased Agonism ◽  
Opioid Ligands ◽  
Substance Abuse Disorders ◽  
Δ Opioid Receptor

Achieving effective pain management is one of the major challenges associated with modern day medicine. Opioids, such as morphine, have been the reference treatment for moderate to severe acute pain not excluding chronic pain modalities. Opioids act through the opioid receptors, the family of G-protein coupled receptors (GPCRs) that mediate pain relief through both the central and peripheral nervous systems. Four types of opioid receptors have been described, including the μ-opioid receptor (MOR), κ-opioid receptor (KOR), δ-opioid receptor (DOR), and the nociceptin opioid peptide receptor (NOP receptor). Despite the proven success of opioids in treating pain, there are still some inherent limitations. All clinically approved MOR analgesics are associated with adverse effects, which include tolerance, dependence, addiction, constipation, and respiratory depression. On the other hand, KOR selective analgesics have found limited clinical utility because they cause sedation, anxiety, dysphoria, and hallucinations. DOR agonists have also been investigated but they have a tendency to cause convulsions. Ligands targeting NOP receptor have been reported in the preclinical literature to be useful as spinal analgesics and as entities against substance abuse disorders while mixed MOR/NOP receptor agonists are useful as analgesics. Ultimately, the goal of opioid-related drug development has always been to design and synthesize derivatives that are equally or more potent than morphine but most importantly are devoid of the dangerous residual side effects and abuse potential. One proposed strategy is to take advantage of biased agonism, in which distinct downstream pathways can be activated by different molecules working through the exact same receptor. It has been proposed that ligands not recruiting β-arrestin 2 or showing a preference for activating a specific G-protein mediated signal transduction pathway will function as safer analgesic across all opioid subtypes. This review will focus on the design and the pharmacological outcomes of biased ligands at the opioid receptors, aiming at achieving functional selectivity.

scholarly journals Rubsicolins are naturally occurring G-protein-biased delta opioid receptor peptides

2018 ◽  
Author(s):  
Robert J. Cassell ◽  
Kendall L. Mores ◽  
Breanna L. Zerfas ◽  
Amr H. Mahmoud ◽  
Markus A. Lill ◽  
...  
Keyword(s):  
G Protein ◽  
Opioid Receptor ◽  
Small Molecule ◽  
G Protein Signaling ◽  
Protein Signaling ◽  
Naturally Occurring ◽  
Δ Opioid Receptor ◽  
The Impact ◽  
G Protein Coupled ◽  
Deltorphin Ii

AbstractThe impact that β-arrestin proteins have on G-protein-coupled receptor trafficking, signaling and physiological behavior has gained much appreciation over the past decade. A number of studies have attributed the side effects associated with the use of naturally occurring and synthetic opioids, such as respiratory depression and constipation, to excessive recruitment of β-arrestin. These findings have led to the development of biased opioid small molecule agonists that do not recruit β-arrestin, activating only the canonical G-protein pathway. Similar G-protein biased small molecule opioids have been found to occur in nature, particularly within kratom, and opioids within salvia have served as a template for the synthesis of other G-protein-biased opioids. Here, we present the first report of naturally occurring peptides that selectively activate G-protein signaling pathways with minimal β-arrestin recruitment. We find that rubiscolin peptides, which are produced as cleavage products of the plant protein rubisco, bind to and activate G-protein signaling at δ opioid receptors. However, unlike the naturally occurring δ opioid peptides leu-enkephalin and deltorphin II, the rubiscolin peptides only very weakly recruit β-arrestin 2 and have undectable recruitment of β-arrestin 1 at the δ opioid receptor.

scholarly journals Discovery of μ, δ-Opioid receptor dual biased agonists that overcome the limitation of prior biased agonists

2021 ◽  
Author(s):  
Jin Hee Lee ◽  
Suh-Youn Shon ◽  
Woojin Jeon ◽  
Sung-Jun Hong ◽  
Junsu Ban ◽  
...  
Keyword(s):  
G Protein ◽  
Opioid Receptor ◽  
Opioid Receptors ◽  
Specific Binding ◽  
Binding Modes ◽  
A Cell ◽  
Μ Opioid Receptor ◽  
Δ Opioid Receptor ◽  
Agonistic Effect ◽  
Biased Agonist

ABSTRACTMorphine is widely used to manage pain in patients, although the risk of side effects is significant. The use of biased agonists to the G protein of μ-opioid receptors has been suggested as a potential solution, although Oliceridine and PZM21 have previously failed to demonstrate benefits in clinical studies. An amplification-induced confusion in the process of comparing G protein and beta-arrestin pathways may account for previous biased agonist mis-identification. Here, we have devised a strategy to discover biased agonists with intrinsic efficacy. We computationally simulated 430,000 molecular dockings to the μ-opioid receptor to construct a compound library. Hits were then verified by experiment. Using the verified compounds, we performed simulations to build a second library with a common scaffold, and selected compounds which show biased features to μ and δ-opioid receptors through a cell-based assay. Three compounds (ID110460001, ID110460002, and ID110460003) with a dual biased agonistic effect for μ and δ-opioid receptors were identified. These candidates are full agonists for the μ-opioid receptor, and they show specific binding modes. Based on our findings, we expect our novel compound to act as a biased agonist than conventional drugs such as Oliceridine.

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