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Agonist-selective recruitment of engineered protein probes and of GRK2 by opioid receptors in living cells
G protein-coupled receptors (GPCRs) signal through allostery, and it is increasingly clear that chemically distinct agonists can produce different receptor-based effects. It has been proposed that agonists selectively promote receptors to recruit one cellular interacting partner over another, introd...
Autores principales: | , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
eLife Sciences Publications, Ltd
2020
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7041944/ https://www.ncbi.nlm.nih.gov/pubmed/32096468 http://dx.doi.org/10.7554/eLife.54208 |
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author | Stoeber, Miriam Jullié, Damien Li, Joy Chakraborty, Soumen Majumdar, Susruta Lambert, Nevin A Manglik, Aashish von Zastrow, Mark |
author_facet | Stoeber, Miriam Jullié, Damien Li, Joy Chakraborty, Soumen Majumdar, Susruta Lambert, Nevin A Manglik, Aashish von Zastrow, Mark |
author_sort | Stoeber, Miriam |
collection | PubMed |
description | G protein-coupled receptors (GPCRs) signal through allostery, and it is increasingly clear that chemically distinct agonists can produce different receptor-based effects. It has been proposed that agonists selectively promote receptors to recruit one cellular interacting partner over another, introducing allosteric ‘bias’ into the signaling system. However, the underlying hypothesis - that different agonists drive GPCRs to engage different cytoplasmic proteins in living cells - remains untested due to the complexity of readouts through which receptor-proximal interactions are typically inferred. We describe a cell-based assay to overcome this challenge, based on GPCR-interacting biosensors that are disconnected from endogenous transduction mechanisms. Focusing on opioid receptors, we directly demonstrate differences between biosensor recruitment produced by chemically distinct opioid ligands in living cells. We then show that selective recruitment applies to GRK2, a biologically relevant GPCR regulator, through discrete interactions of GRK2 with receptors or with G protein beta-gamma subunits which are differentially promoted by agonists. |
format | Online Article Text |
id | pubmed-7041944 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2020 |
publisher | eLife Sciences Publications, Ltd |
record_format | MEDLINE/PubMed |
spelling | pubmed-70419442020-02-27 Agonist-selective recruitment of engineered protein probes and of GRK2 by opioid receptors in living cells Stoeber, Miriam Jullié, Damien Li, Joy Chakraborty, Soumen Majumdar, Susruta Lambert, Nevin A Manglik, Aashish von Zastrow, Mark eLife Biochemistry and Chemical Biology G protein-coupled receptors (GPCRs) signal through allostery, and it is increasingly clear that chemically distinct agonists can produce different receptor-based effects. It has been proposed that agonists selectively promote receptors to recruit one cellular interacting partner over another, introducing allosteric ‘bias’ into the signaling system. However, the underlying hypothesis - that different agonists drive GPCRs to engage different cytoplasmic proteins in living cells - remains untested due to the complexity of readouts through which receptor-proximal interactions are typically inferred. We describe a cell-based assay to overcome this challenge, based on GPCR-interacting biosensors that are disconnected from endogenous transduction mechanisms. Focusing on opioid receptors, we directly demonstrate differences between biosensor recruitment produced by chemically distinct opioid ligands in living cells. We then show that selective recruitment applies to GRK2, a biologically relevant GPCR regulator, through discrete interactions of GRK2 with receptors or with G protein beta-gamma subunits which are differentially promoted by agonists. eLife Sciences Publications, Ltd 2020-02-25 /pmc/articles/PMC7041944/ /pubmed/32096468 http://dx.doi.org/10.7554/eLife.54208 Text en © 2020, Stoeber et al http://creativecommons.org/licenses/by/4.0/ http://creativecommons.org/licenses/by/4.0/This article is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0/) , which permits unrestricted use and redistribution provided that the original author and source are credited. |
spellingShingle | Biochemistry and Chemical Biology Stoeber, Miriam Jullié, Damien Li, Joy Chakraborty, Soumen Majumdar, Susruta Lambert, Nevin A Manglik, Aashish von Zastrow, Mark Agonist-selective recruitment of engineered protein probes and of GRK2 by opioid receptors in living cells |
title | Agonist-selective recruitment of engineered protein probes and of GRK2 by opioid receptors in living cells |
title_full | Agonist-selective recruitment of engineered protein probes and of GRK2 by opioid receptors in living cells |
title_fullStr | Agonist-selective recruitment of engineered protein probes and of GRK2 by opioid receptors in living cells |
title_full_unstemmed | Agonist-selective recruitment of engineered protein probes and of GRK2 by opioid receptors in living cells |
title_short | Agonist-selective recruitment of engineered protein probes and of GRK2 by opioid receptors in living cells |
title_sort | agonist-selective recruitment of engineered protein probes and of grk2 by opioid receptors in living cells |
topic | Biochemistry and Chemical Biology |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7041944/ https://www.ncbi.nlm.nih.gov/pubmed/32096468 http://dx.doi.org/10.7554/eLife.54208 |
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