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Agonist-induced phosphorylation bar code and differential post-activation signaling of the delta opioid receptor revealed by phosphosite-specific antibodies

The δ-opioid receptor (DOP) is an attractive pharmacological target due to its potent analgesic, anxiolytic and anti-depressant activity in chronic pain models. However, some but not all selective DOP agonists also produce severe adverse effects such as seizures. Thus, the development of novel agoni...

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Autores principales: Mann, Anika, Liebetrau, Sophia, Klima, Marie, Dasgupta, Pooja, Massotte, Dominique, Schulz, Stefan
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Nature Publishing Group UK 2020
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7244497/
https://www.ncbi.nlm.nih.gov/pubmed/32444688
http://dx.doi.org/10.1038/s41598-020-65589-7
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author Mann, Anika
Liebetrau, Sophia
Klima, Marie
Dasgupta, Pooja
Massotte, Dominique
Schulz, Stefan
author_facet Mann, Anika
Liebetrau, Sophia
Klima, Marie
Dasgupta, Pooja
Massotte, Dominique
Schulz, Stefan
author_sort Mann, Anika
collection PubMed
description The δ-opioid receptor (DOP) is an attractive pharmacological target due to its potent analgesic, anxiolytic and anti-depressant activity in chronic pain models. However, some but not all selective DOP agonists also produce severe adverse effects such as seizures. Thus, the development of novel agonists requires a profound understanding of their effects on DOP phosphorylation, post-activation signaling and dephosphorylation. Here we show that agonist-induced DOP phosphorylation at threonine 361 (T361) and serine 363 (S363) proceeds with a temporal hierarchy, with S363 as primary site of phosphorylation. This phosphorylation is mediated by G protein-coupled receptor kinases 2 and 3 (GRK2/3) followed by DOP endocytosis and desensitization. DOP dephosphorylation occurs within minutes and is predominantly mediated by protein phosphatases (PP) 1α and 1β. A comparison of structurally diverse DOP agonists and clinically used opioids demonstrated high correlation between G protein-dependent signaling efficacies and receptor internalization. In vivo, DOP agonists induce receptor phosphorylation in a dose-dependent and agonist-selective manner that could be blocked by naltrexone in DOP-eGFP mice. Together, our studies provide novel tools and insights for ligand-activated DOP signaling in vitro and in vivo and suggest that DOP agonist efficacies may determine receptor post-activation signaling.
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spelling pubmed-72444972020-05-30 Agonist-induced phosphorylation bar code and differential post-activation signaling of the delta opioid receptor revealed by phosphosite-specific antibodies Mann, Anika Liebetrau, Sophia Klima, Marie Dasgupta, Pooja Massotte, Dominique Schulz, Stefan Sci Rep Article The δ-opioid receptor (DOP) is an attractive pharmacological target due to its potent analgesic, anxiolytic and anti-depressant activity in chronic pain models. However, some but not all selective DOP agonists also produce severe adverse effects such as seizures. Thus, the development of novel agonists requires a profound understanding of their effects on DOP phosphorylation, post-activation signaling and dephosphorylation. Here we show that agonist-induced DOP phosphorylation at threonine 361 (T361) and serine 363 (S363) proceeds with a temporal hierarchy, with S363 as primary site of phosphorylation. This phosphorylation is mediated by G protein-coupled receptor kinases 2 and 3 (GRK2/3) followed by DOP endocytosis and desensitization. DOP dephosphorylation occurs within minutes and is predominantly mediated by protein phosphatases (PP) 1α and 1β. A comparison of structurally diverse DOP agonists and clinically used opioids demonstrated high correlation between G protein-dependent signaling efficacies and receptor internalization. In vivo, DOP agonists induce receptor phosphorylation in a dose-dependent and agonist-selective manner that could be blocked by naltrexone in DOP-eGFP mice. Together, our studies provide novel tools and insights for ligand-activated DOP signaling in vitro and in vivo and suggest that DOP agonist efficacies may determine receptor post-activation signaling. Nature Publishing Group UK 2020-05-22 /pmc/articles/PMC7244497/ /pubmed/32444688 http://dx.doi.org/10.1038/s41598-020-65589-7 Text en © The Author(s) 2020 Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.
spellingShingle Article
Mann, Anika
Liebetrau, Sophia
Klima, Marie
Dasgupta, Pooja
Massotte, Dominique
Schulz, Stefan
Agonist-induced phosphorylation bar code and differential post-activation signaling of the delta opioid receptor revealed by phosphosite-specific antibodies
title Agonist-induced phosphorylation bar code and differential post-activation signaling of the delta opioid receptor revealed by phosphosite-specific antibodies
title_full Agonist-induced phosphorylation bar code and differential post-activation signaling of the delta opioid receptor revealed by phosphosite-specific antibodies
title_fullStr Agonist-induced phosphorylation bar code and differential post-activation signaling of the delta opioid receptor revealed by phosphosite-specific antibodies
title_full_unstemmed Agonist-induced phosphorylation bar code and differential post-activation signaling of the delta opioid receptor revealed by phosphosite-specific antibodies
title_short Agonist-induced phosphorylation bar code and differential post-activation signaling of the delta opioid receptor revealed by phosphosite-specific antibodies
title_sort agonist-induced phosphorylation bar code and differential post-activation signaling of the delta opioid receptor revealed by phosphosite-specific antibodies
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7244497/
https://www.ncbi.nlm.nih.gov/pubmed/32444688
http://dx.doi.org/10.1038/s41598-020-65589-7
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