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The GPCR-associated sorting protein 1 regulates ligand-induced down-regulation of GPR55.
Br J Pharmacol. 2012 Apr; 165(8):2611-9.BJ

Abstract

BACKGROUND AND PURPOSE

Many GPCRs, including the CB(1) cannabinoid receptor, are down-regulated following prolonged agonist exposure by interacting with the GPCR-associated sorting protein-1 (GASP-1). The CB(1) receptor antagonist rimonabant has also recently been described to be an agonist at GPR55, a cannabinoid-related receptor. Here we investigated the post-endocytic properties of GPR55 after agonist exposure and tested whether GASP-1 is involved in this process.

EXPERIMENTAL APPROACH

We evaluated the direct protein-protein interaction of GPR55 with GASP-1 using (i) GST-binding assays and (ii) co-immunoprecipitation assays in GPR55-HEK293 cells with endogenous GASP-1 expression. We further tested the internalization, recycling and degradation of GPR55 using confocal fluorescence microscopy and biotinylation assays in the presence and absence of GASP-1 (lentiviral small hairpin RNA knockdown of GASP-1) under prolonged agonist [rimonabant (RIM), lysophosphatidylinositol (LPI)] stimulation.

KEY RESULTS

We showed that the prolonged activation of GPR55 with rimonabant or LPI down-regulates GPR55 via GASP-1. GASP-1 binds to GPR55 in vitro, and this interaction was required for targeting GPR55 for degradation. Disrupting the GPR55-GASP-1 interaction prevented post-endocytic receptor degradation, and thereby allowed receptor recycling.

CONCLUSION AND IMPLICATIONS

These data implicate GASP-1 as an important regulator of ligand-mediated down-regulation of GPR55. By identifying GASP-1 as a key regulator of the trafficking and, by extension, functional expression of GPR55, we may be one step closer to gaining a better understanding of this receptor in response to cannabinoid drugs.

LINKED ARTICLES

This article is part of a themed section on Cannabinoids in Biology and Medicine. To view the other articles in this section visit http://dx.doi.org/10.1111/bph.2012.165.issue-8. To view Part I of Cannabinoids in Biology and Medicine visit http://dx.doi.org/10.1111/bph.2011.163.issue-7.

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Authors+Show Affiliations

Kargl J
Institute for Experimental and Clinical Pharmacology, Medical University of Graz, Graz, Austria. julia.kargl@medunigraz.at
Balenga NA
No affiliation info available
Platzer W
No affiliation info available
Martini L
No affiliation info available
Whistler JL
No affiliation info available
Waldhoer M
No affiliation info available

MeSH

Down-RegulationGlutathione TransferaseHEK293 CellsHumansLigandsLysophospholipidsPiperidinesPyrazolesReceptor, Cannabinoid, CB1Receptors, CannabinoidReceptors, G-Protein-CoupledRecombinant Fusion ProteinsRimonabantVesicular Transport Proteins

Pub Type(s)

Journal Article
Research Support, Non-U.S. Gov't

Language

eng

PubMed ID

21718301

Citation

Kargl, J, et al. "The GPCR-associated Sorting Protein 1 Regulates Ligand-induced Down-regulation of GPR55." British Journal of Pharmacology, vol. 165, no. 8, 2012, pp. 2611-9.
Kargl J, Balenga NA, Platzer W, et al. The GPCR-associated sorting protein 1 regulates ligand-induced down-regulation of GPR55. Br J Pharmacol. 2012;165(8):2611-9.
Kargl, J., Balenga, N. A., Platzer, W., Martini, L., Whistler, J. L., & Waldhoer, M. (2012). The GPCR-associated sorting protein 1 regulates ligand-induced down-regulation of GPR55. British Journal of Pharmacology, 165(8), 2611-9. https://doi.org/10.1111/j.1476-5381.2011.01562.x
Kargl J, et al. The GPCR-associated Sorting Protein 1 Regulates Ligand-induced Down-regulation of GPR55. Br J Pharmacol. 2012;165(8):2611-9. PubMed PMID: 21718301.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - The GPCR-associated sorting protein 1 regulates ligand-induced down-regulation of GPR55. AU - Kargl,J, AU - Balenga,N A, AU - Platzer,W, AU - Martini,L, AU - Whistler,J L, AU - Waldhoer,M, PY - 2011/7/2/entrez PY - 2011/7/2/pubmed PY - 2012/7/28/medline SP - 2611 EP - 9 JF - British journal of pharmacology JO - Br J Pharmacol VL - 165 IS - 8 N2 - BACKGROUND AND PURPOSE: Many GPCRs, including the CB(1) cannabinoid receptor, are down-regulated following prolonged agonist exposure by interacting with the GPCR-associated sorting protein-1 (GASP-1). The CB(1) receptor antagonist rimonabant has also recently been described to be an agonist at GPR55, a cannabinoid-related receptor. Here we investigated the post-endocytic properties of GPR55 after agonist exposure and tested whether GASP-1 is involved in this process. EXPERIMENTAL APPROACH: We evaluated the direct protein-protein interaction of GPR55 with GASP-1 using (i) GST-binding assays and (ii) co-immunoprecipitation assays in GPR55-HEK293 cells with endogenous GASP-1 expression. We further tested the internalization, recycling and degradation of GPR55 using confocal fluorescence microscopy and biotinylation assays in the presence and absence of GASP-1 (lentiviral small hairpin RNA knockdown of GASP-1) under prolonged agonist [rimonabant (RIM), lysophosphatidylinositol (LPI)] stimulation. KEY RESULTS: We showed that the prolonged activation of GPR55 with rimonabant or LPI down-regulates GPR55 via GASP-1. GASP-1 binds to GPR55 in vitro, and this interaction was required for targeting GPR55 for degradation. Disrupting the GPR55-GASP-1 interaction prevented post-endocytic receptor degradation, and thereby allowed receptor recycling. CONCLUSION AND IMPLICATIONS: These data implicate GASP-1 as an important regulator of ligand-mediated down-regulation of GPR55. By identifying GASP-1 as a key regulator of the trafficking and, by extension, functional expression of GPR55, we may be one step closer to gaining a better understanding of this receptor in response to cannabinoid drugs. LINKED ARTICLES: This article is part of a themed section on Cannabinoids in Biology and Medicine. To view the other articles in this section visit http://dx.doi.org/10.1111/bph.2012.165.issue-8. To view Part I of Cannabinoids in Biology and Medicine visit http://dx.doi.org/10.1111/bph.2011.163.issue-7. SN - 1476-5381 UR - https://www.unboundmedicine.com/medline/citation/21718301/The_GPCR_associated_sorting_protein_1_regulates_ligand_induced_down_regulation_of_GPR55_ L2 - https://doi.org/10.1111/j.1476-5381.2011.01562.x DB - PRIME DP - Unbound Medicine ER -