Delineating the interactions between the cannabinoid CB2 receptor and its regulatory effectors; β-arrestins and G protein-coupled receptor kinases

Patel, Monica; Matti, Christoph; Grimsey, Natasha L.; Legler, Daniel F.; Javitch, Jonathan A.; Finlay, David B.; Glass, Michelle

Delineating the interactions between the cannabinoid CB2 receptor and its regulatory effectors; β-arrestins and G protein-coupled receptor kinases

Dateien

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Datum

2022

Autor:innen

Patel, Monica

Matti, Christoph

Grimsey, Natasha L.

Legler, Daniel F.

Javitch, Jonathan A.

Finlay, David B.

Glass, Michelle

Publikationstyp

Zeitschriftenartikel

Publikationsstatus

Published

Erschienen in

British Journal of Pharmacology (BJP). Wiley. 2022, 179(10), pp. 2223-2239. ISSN 0366-0826. eISSN 1476-5381. Available under: doi: 10.1111/bph.15748

Zusammenfassung

Background and Purpose
The cannabinoid CB₂ receptor (CB₂) is a promising therapeutic target for modulating inflammation. However, little is known surrounding the mechanisms underpinning CB₂ desensitisation and regulation, particularly the role of G protein-coupled receptor kinases (GRKs). Here, we evaluated the role of six GRK isoforms in β-arrestin recruitment to CB₂. Mutagenesis of several distal C-terminal aspartic acid residues was also performed in an attempt to delineate additional structural elements involved in the regulation of CB₂.

Experimental Approach
In CB₂-expressing HEK 293 cells, β-arrestin translocation was measured using real-time BRET assays. G protein dissociation BRET assays were performed to assess the activation and desensitisation of CB₂ in the presence of β-arrestin 2.

Key Results
Overexpression of GRK isoforms 1-6 failed to considerably improve translocation of either β-arrestin 1 or β-arrestin 2 to CB₂. Consistent with this, inhibition of endogenous GRK2/3 did not substantially reduce β-arrestin 2 translocation. Mutagenesis of C-terminal aspartic acid residues resulted in attenuation of β-arrestin 2 translocation, which translated to a reduction in desensitisation of G protein activation.

Conclusion and Implications
Our findings suggest that CB₂ does not adhere to the classical GPCR regulatory paradigm, entailing GRK- and β-arrestin-mediated desensitisation. Instead, C-terminal aspartic acid residues may act as phospho-mimics to induce β-arrestin activation. This study provides novel insights into the regulatory mechanisms of CB₂, which may aid in our understanding of drug tolerance and dependence.

Fachgebiet (DDC)

570 Biowissenschaften, Biologie

Schlagwörter

Cannabinoid CB2 receptor, β-arrestin, G protein-coupled receptor kinase

Zitieren

ISO 690

PATEL, Monica, Christoph MATTI, Natasha L. GRIMSEY, Daniel F. LEGLER, Jonathan A. JAVITCH, David B. FINLAY, Michelle GLASS, 2022. Delineating the interactions between the cannabinoid CB2 receptor and its regulatory effectors; β-arrestins and G protein-coupled receptor kinases. In: British Journal of Pharmacology (BJP). Wiley. 2022, 179(10), pp. 2223-2239. ISSN 0366-0826. eISSN 1476-5381. Available under: doi: 10.1111/bph.15748

BibTex

@article{Patel2022-05Delin-55750,
  year={2022},
  doi={10.1111/bph.15748},
  title={Delineating the interactions between the cannabinoid CB2 receptor and its regulatory effectors; β-arrestins and G protein-coupled receptor kinases},
  number={10},
  volume={179},
  issn={0366-0826},
  journal={British Journal of Pharmacology (BJP)},
  pages={2223--2239},
  author={Patel, Monica and Matti, Christoph and Grimsey, Natasha L. and Legler, Daniel F. and Javitch, Jonathan A. and Finlay, David B. and Glass, Michelle}
}

RDF

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    <dcterms:abstract xml:lang="eng">Background and Purpose&lt;br /&gt;The cannabinoid CB&lt;sub&gt;2&lt;/sub&gt; receptor (CB&lt;sub&gt;2&lt;/sub&gt;) is a promising therapeutic target for modulating inflammation. However, little is known surrounding the mechanisms underpinning CB&lt;sub&gt;2&lt;/sub&gt; desensitisation and regulation, particularly the role of G protein-coupled receptor kinases (GRKs). Here, we evaluated the role of six GRK isoforms in β-arrestin recruitment to CB&lt;sub&gt;2&lt;/sub&gt;. Mutagenesis of several distal C-terminal aspartic acid residues was also performed in an attempt to delineate additional structural elements involved in the regulation of CB&lt;sub&gt;2&lt;/sub&gt;.&lt;br /&gt;&lt;br /&gt;Experimental Approach&lt;br /&gt;In CB&lt;sub&gt;2&lt;/sub&gt;-expressing HEK 293 cells, β-arrestin translocation was measured using real-time BRET assays. G protein dissociation BRET assays were performed to assess the activation and desensitisation of CB&lt;sub&gt;2&lt;/sub&gt; in the presence of β-arrestin 2.&lt;br /&gt;&lt;br /&gt;Key Results&lt;br /&gt;Overexpression of GRK isoforms 1-6 failed to considerably improve translocation of either β-arrestin 1 or β-arrestin 2 to CB&lt;sub&gt;2&lt;/sub&gt;. Consistent with this, inhibition of endogenous GRK2/3 did not substantially reduce β-arrestin 2 translocation. Mutagenesis of C-terminal aspartic acid residues resulted in attenuation of β-arrestin 2 translocation, which translated to a reduction in desensitisation of G protein activation.&lt;br /&gt;&lt;br /&gt;Conclusion and Implications&lt;br /&gt;Our findings suggest that CB&lt;sub&gt;2&lt;/sub&gt; does not adhere to the classical GPCR regulatory paradigm, entailing GRK- and β-arrestin-mediated desensitisation. Instead, C-terminal aspartic acid residues may act as phospho-mimics to induce β-arrestin activation. This study provides novel insights into the regulatory mechanisms of CB&lt;sub&gt;2&lt;/sub&gt;, which may aid in our understanding of drug tolerance and dependence.</dcterms:abstract>
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