New phosphosite-specific antibodies to unravel the role of GRK phosphorylation in dopamine D2 receptor regulation and signaling

Abstract The dopamine D2 receptor (D2R) is the target of drugs used to treat the symptoms of Parkinson’s disease and schizophrenia. The D2R is regulated through its interaction with and phosphorylation by G protein receptor kinases (GRKs) and interaction with arrestins. More recently, D2R arrestin-m...

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Autores principales:	Anika Mann, Alastair C. Keen, Hanka Mark, Pooja Dasgupta, Jonathan A. Javitch, Meritxell Canals, Stefan Schulz, J. Robert Lane
Formato:	article
Lenguaje:	EN
Publicado:	Nature Portfolio 2021
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/ca749480bcc9489bb045eb4ed1f42bf3
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spelling	oai:doaj.org-article:ca749480bcc9489bb045eb4ed1f42bf32021-12-02T18:03:32ZNew phosphosite-specific antibodies to unravel the role of GRK phosphorylation in dopamine D2 receptor regulation and signaling10.1038/s41598-021-87417-22045-2322https://doaj.org/article/ca749480bcc9489bb045eb4ed1f42bf32021-04-01T00:00:00Zhttps://doi.org/10.1038/s41598-021-87417-2https://doaj.org/toc/2045-2322Abstract The dopamine D2 receptor (D2R) is the target of drugs used to treat the symptoms of Parkinson’s disease and schizophrenia. The D2R is regulated through its interaction with and phosphorylation by G protein receptor kinases (GRKs) and interaction with arrestins. More recently, D2R arrestin-mediated signaling has been shown to have distinct physiological functions to those of G protein signalling. Relatively little is known regarding the patterns of D2R phosphorylation that might control these processes. We aimed to generate antibodies specific for intracellular D2R phosphorylation sites to facilitate the investigation of these mechanisms. We synthesised double phosphorylated peptides corresponding to regions within intracellular loop 3 of the hD2R and used them to raise phosphosite-specific antibodies to capture a broad screen of GRK-mediated phosphorylation. We identify an antibody specific to a GRK2/3 phosphorylation site in intracellular loop 3 of the D2R. We compared measurements of D2R phosphorylation with other measurements of D2R signalling to profile selected D2R agonists including previously described biased agonists. These studies demonstrate the utility of novel phosphosite-specific antibodies to investigate D2R regulation and signalling.Anika MannAlastair C. KeenHanka MarkPooja DasguptaJonathan A. JavitchMeritxell CanalsStefan SchulzJ. Robert LaneNature PortfolioarticleMedicineRScienceQENScientific Reports, Vol 11, Iss 1, Pp 1-17 (2021)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Anika Mann Alastair C. Keen Hanka Mark Pooja Dasgupta Jonathan A. Javitch Meritxell Canals Stefan Schulz J. Robert Lane New phosphosite-specific antibodies to unravel the role of GRK phosphorylation in dopamine D2 receptor regulation and signaling
description	Abstract The dopamine D2 receptor (D2R) is the target of drugs used to treat the symptoms of Parkinson’s disease and schizophrenia. The D2R is regulated through its interaction with and phosphorylation by G protein receptor kinases (GRKs) and interaction with arrestins. More recently, D2R arrestin-mediated signaling has been shown to have distinct physiological functions to those of G protein signalling. Relatively little is known regarding the patterns of D2R phosphorylation that might control these processes. We aimed to generate antibodies specific for intracellular D2R phosphorylation sites to facilitate the investigation of these mechanisms. We synthesised double phosphorylated peptides corresponding to regions within intracellular loop 3 of the hD2R and used them to raise phosphosite-specific antibodies to capture a broad screen of GRK-mediated phosphorylation. We identify an antibody specific to a GRK2/3 phosphorylation site in intracellular loop 3 of the D2R. We compared measurements of D2R phosphorylation with other measurements of D2R signalling to profile selected D2R agonists including previously described biased agonists. These studies demonstrate the utility of novel phosphosite-specific antibodies to investigate D2R regulation and signalling.
format	article
author	Anika Mann Alastair C. Keen Hanka Mark Pooja Dasgupta Jonathan A. Javitch Meritxell Canals Stefan Schulz J. Robert Lane
author_facet	Anika Mann Alastair C. Keen Hanka Mark Pooja Dasgupta Jonathan A. Javitch Meritxell Canals Stefan Schulz J. Robert Lane
author_sort	Anika Mann
title	New phosphosite-specific antibodies to unravel the role of GRK phosphorylation in dopamine D2 receptor regulation and signaling
title_short	New phosphosite-specific antibodies to unravel the role of GRK phosphorylation in dopamine D2 receptor regulation and signaling
title_full	New phosphosite-specific antibodies to unravel the role of GRK phosphorylation in dopamine D2 receptor regulation and signaling
title_fullStr	New phosphosite-specific antibodies to unravel the role of GRK phosphorylation in dopamine D2 receptor regulation and signaling
title_full_unstemmed	New phosphosite-specific antibodies to unravel the role of GRK phosphorylation in dopamine D2 receptor regulation and signaling
title_sort	new phosphosite-specific antibodies to unravel the role of grk phosphorylation in dopamine d2 receptor regulation and signaling
publisher	Nature Portfolio
publishDate	2021
url	https://doaj.org/article/ca749480bcc9489bb045eb4ed1f42bf3
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New phosphosite-specific antibodies to unravel the role of GRK phosphorylation in dopamine D2 receptor regulation and signaling

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