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Ligand-Independent G Protein–Coupled Estrogen Receptor/G Protein–Coupled Receptor 30 Activity: Lack of Receptor-Dependent Effects of G-1 and 17β-Estradiol

G protein–coupled receptor 30 (GPR30) is a membrane receptor reported to bind 17β-estradiol (E2) and mediate rapid nongenomic estrogen responses, hence also named G protein–coupled estrogen receptor. G-1 is a proposed GPR30-specific agonist that has been used to implicate the receptor in several pat...

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Autores principales: Tutzauer, Julia, Gonzalez de Valdivia, Ernesto, Swärd, Karl, Alexandrakis Eilard, Ioannis, Broselid, Stefan, Kahn, Robin, Olde, Björn, Leeb-Lundberg, L. M. Fredrik
Formato: Online Artículo Texto
Lenguaje:English
Publicado: The American Society for Pharmacology and Experimental Therapeutics 2021
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8626787/
https://www.ncbi.nlm.nih.gov/pubmed/34330822
http://dx.doi.org/10.1124/molpharm.121.000259
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author Tutzauer, Julia
Gonzalez de Valdivia, Ernesto
Swärd, Karl
Alexandrakis Eilard, Ioannis
Broselid, Stefan
Kahn, Robin
Olde, Björn
Leeb-Lundberg, L. M. Fredrik
author_facet Tutzauer, Julia
Gonzalez de Valdivia, Ernesto
Swärd, Karl
Alexandrakis Eilard, Ioannis
Broselid, Stefan
Kahn, Robin
Olde, Björn
Leeb-Lundberg, L. M. Fredrik
author_sort Tutzauer, Julia
collection PubMed
description G protein–coupled receptor 30 (GPR30) is a membrane receptor reported to bind 17β-estradiol (E2) and mediate rapid nongenomic estrogen responses, hence also named G protein–coupled estrogen receptor. G-1 is a proposed GPR30-specific agonist that has been used to implicate the receptor in several pathophysiological events. However, controversy surrounds the role of GPR30 in G-1 and E2 responses. We investigated GPR30 activity in the absence and presence of G-1 and E2 in several eukaryotic systems ex vivo and in vitro in the absence and presence of the receptor. Ex vivo activity was addressed using the caudal artery from wild-type (WT) and GPR30 knockout (KO) mice, and in vitro activity was addressed using a HeLa cell line stably expressing a synthetic multifunctional promoter (nuclear factor κB, signal transducer and activator of transcription, activator protein 1)–luciferase construct (HFF11 cells) and a human GPR30-inducible T-REx system (T-REx HFF11 cells), HFF11 and human embryonic kidney 293 cells transiently expressing WT GPR30 and GPR30 lacking the C-terminal PDZ (postsynaptic density-95/discs-large /zonula occludens-1 homology) motif SSAV, and yeast Saccharomyces cerevisiae transformed to express GPR30. WT and KO arteries exhibited similar contractile responses to 60 mM KCl and 0.3 μM cirazoline, and G-1 relaxed both arteries with the same potency and efficacy. Furthermore, expression of GPR30 did not introduce any responses to 1 μM G-1 and 0.1 μM E2 in vitro. On the other hand, receptor expression caused considerable ligand-independent activity in vitro, which was receptor PDZ motif-dependent in mammalian cells. We conclude from these results that GPR30 exhibits ligand-independent activity in vitro but no G-1– or E2-stimulated activity in any of the systems used. SIGNIFICANCE STATEMENT: Much controversy surrounds 17β-estradiol (E2) and G-1 as G protein–coupled receptor 30 (GPR30) agonists. We used several recombinant eukaryotic systems ex vivo and in vitro with and without GPR30 expression to address the role of this receptor in responses to these proposed agonists. Our results show that GPR30 exhibits considerable ligand-independent activity in vitro but no G-1– or E2-stimulated activity in any of the systems used. Thus, classifying GPR30 as an estrogen receptor and G-1 as a specific GPR30 agonist is unfounded.
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spelling pubmed-86267872022-09-01 Ligand-Independent G Protein–Coupled Estrogen Receptor/G Protein–Coupled Receptor 30 Activity: Lack of Receptor-Dependent Effects of G-1 and 17β-Estradiol Tutzauer, Julia Gonzalez de Valdivia, Ernesto Swärd, Karl Alexandrakis Eilard, Ioannis Broselid, Stefan Kahn, Robin Olde, Björn Leeb-Lundberg, L. M. Fredrik Mol Pharmacol Articles G protein–coupled receptor 30 (GPR30) is a membrane receptor reported to bind 17β-estradiol (E2) and mediate rapid nongenomic estrogen responses, hence also named G protein–coupled estrogen receptor. G-1 is a proposed GPR30-specific agonist that has been used to implicate the receptor in several pathophysiological events. However, controversy surrounds the role of GPR30 in G-1 and E2 responses. We investigated GPR30 activity in the absence and presence of G-1 and E2 in several eukaryotic systems ex vivo and in vitro in the absence and presence of the receptor. Ex vivo activity was addressed using the caudal artery from wild-type (WT) and GPR30 knockout (KO) mice, and in vitro activity was addressed using a HeLa cell line stably expressing a synthetic multifunctional promoter (nuclear factor κB, signal transducer and activator of transcription, activator protein 1)–luciferase construct (HFF11 cells) and a human GPR30-inducible T-REx system (T-REx HFF11 cells), HFF11 and human embryonic kidney 293 cells transiently expressing WT GPR30 and GPR30 lacking the C-terminal PDZ (postsynaptic density-95/discs-large /zonula occludens-1 homology) motif SSAV, and yeast Saccharomyces cerevisiae transformed to express GPR30. WT and KO arteries exhibited similar contractile responses to 60 mM KCl and 0.3 μM cirazoline, and G-1 relaxed both arteries with the same potency and efficacy. Furthermore, expression of GPR30 did not introduce any responses to 1 μM G-1 and 0.1 μM E2 in vitro. On the other hand, receptor expression caused considerable ligand-independent activity in vitro, which was receptor PDZ motif-dependent in mammalian cells. We conclude from these results that GPR30 exhibits ligand-independent activity in vitro but no G-1– or E2-stimulated activity in any of the systems used. SIGNIFICANCE STATEMENT: Much controversy surrounds 17β-estradiol (E2) and G-1 as G protein–coupled receptor 30 (GPR30) agonists. We used several recombinant eukaryotic systems ex vivo and in vitro with and without GPR30 expression to address the role of this receptor in responses to these proposed agonists. Our results show that GPR30 exhibits considerable ligand-independent activity in vitro but no G-1– or E2-stimulated activity in any of the systems used. Thus, classifying GPR30 as an estrogen receptor and G-1 as a specific GPR30 agonist is unfounded. The American Society for Pharmacology and Experimental Therapeutics 2021-09 2021-09 /pmc/articles/PMC8626787/ /pubmed/34330822 http://dx.doi.org/10.1124/molpharm.121.000259 Text en Copyright © 2021 by The Author(s) https://creativecommons.org/licenses/by-nc/4.0/This is an open access article distributed under the CC BY-NC Attribution 4.0 International license (https://creativecommons.org/licenses/by-nc/4.0/) .
spellingShingle Articles
Tutzauer, Julia
Gonzalez de Valdivia, Ernesto
Swärd, Karl
Alexandrakis Eilard, Ioannis
Broselid, Stefan
Kahn, Robin
Olde, Björn
Leeb-Lundberg, L. M. Fredrik
Ligand-Independent G Protein–Coupled Estrogen Receptor/G Protein–Coupled Receptor 30 Activity: Lack of Receptor-Dependent Effects of G-1 and 17β-Estradiol
title Ligand-Independent G Protein–Coupled Estrogen Receptor/G Protein–Coupled Receptor 30 Activity: Lack of Receptor-Dependent Effects of G-1 and 17β-Estradiol
title_full Ligand-Independent G Protein–Coupled Estrogen Receptor/G Protein–Coupled Receptor 30 Activity: Lack of Receptor-Dependent Effects of G-1 and 17β-Estradiol
title_fullStr Ligand-Independent G Protein–Coupled Estrogen Receptor/G Protein–Coupled Receptor 30 Activity: Lack of Receptor-Dependent Effects of G-1 and 17β-Estradiol
title_full_unstemmed Ligand-Independent G Protein–Coupled Estrogen Receptor/G Protein–Coupled Receptor 30 Activity: Lack of Receptor-Dependent Effects of G-1 and 17β-Estradiol
title_short Ligand-Independent G Protein–Coupled Estrogen Receptor/G Protein–Coupled Receptor 30 Activity: Lack of Receptor-Dependent Effects of G-1 and 17β-Estradiol
title_sort ligand-independent g protein–coupled estrogen receptor/g protein–coupled receptor 30 activity: lack of receptor-dependent effects of g-1 and 17β-estradiol
topic Articles
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8626787/
https://www.ncbi.nlm.nih.gov/pubmed/34330822
http://dx.doi.org/10.1124/molpharm.121.000259
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