Bibliographic Details
| Title: |
SNAP-tag-enabled super-resolution imaging reveals constitutive and agonist-dependent trafficking of GPR56 in pancreatic β-cells |
| Authors: |
Oladapo E. Olaniru, Jordan Cheng, Julia Ast, Anastasia Arvaniti, Patricio Atanes, Guo C. Huang, Aileen J.F. King, Peter M. Jones, Johannes Broichhagen, David J. Hodson, Shanta J. Persaud |
| Source: |
Molecular Metabolism, Vol 53, Iss , Pp 101285- (2021) |
| Publisher Information: |
Elsevier, 2021. |
| Publication Year: |
2021 |
| Collection: |
LCC:Internal medicine |
| Subject Terms: |
GPR56, SNAP-tag, Trafficking, Islets, Apoptosis, CRISPR-Cas9, Internal medicine, RC31-1245 |
| More Details: |
Objective: Members of the adhesion G protein-coupled receptor (aGPCR) subfamily are important actors in metabolic processes, with GPR56 (ADGRG1) emerging as a possible target for type 2 diabetes therapy. GPR56 can be activated by collagen III, its endogenous ligand, and by a synthetic seven amino-acid peptide (TYFAVLM; P7) contained within the GPR56 Stachel sequence. However, the mechanisms regulating GPR56 trafficking dynamics and agonist activities are not yet clear. Methods: Here, we introduced SNAPf-tag into the N-terminal segment of GPR56 to monitor GPR56 cellular activity in situ. Confocal and super-resolution microscopy were used to investigate the trafficking pattern of GPR56 in native MIN6 β-cells and in MIN6 β-cells where GPR56 had been deleted by CRISPR-Cas9 gene editing. Insulin secretion, changes in intracellular calcium, and β-cell apoptosis were determined by radioimmunoassay, single-cell calcium microfluorimetry, and measuring caspase 3/7 activities, respectively, in MIN6 β-cells and human islets. Results: SNAP-tag labelling indicated that GPR56 predominantly underwent constitutive internalisation in the absence of an exogenous agonist, unlike GLP-1R. Collagen III further stimulated GPR56 internalisation, whereas P7 was without significant effect. The overexpression of GPR56 in MIN6 β-cells did not affect insulin secretion. However, it was associated with reduced β-cell apoptosis, while the deletion of GPR56 made MIN6 β-cells more susceptible to cytokine-induced apoptosis. P7 induced a rapid increase in the intracellular calcium in MIN6 β-cells (in a GPR56-dependent manner) and human islets, and it also caused a sustained and reversible increase in insulin secretion from human islets. Collagen III protected human islets from cytokine-induced apoptosis, while P7 was without significant effect. Conclusions: These data indicate that GPR56 exhibits both agonist-dependent and -independent trafficking in β-cells and suggest that while GPR56 undergoes constitutive signalling, it can also respond to its ligands when required. We have also identified that constitutive and agonist-dependent GPR56 activation is coupled to protect β-cells against apoptosis, offering a potential therapeutic target to maintain β-cell mass in type 2 diabetes. |
| Document Type: |
article |
| File Description: |
electronic resource |
| Language: |
English |
| ISSN: |
2212-8778 |
| Relation: |
http://www.sciencedirect.com/science/article/pii/S2212877821001307; https://doaj.org/toc/2212-8778 |
| DOI: |
10.1016/j.molmet.2021.101285 |
| Access URL: |
https://doaj.org/article/3df09aa019eb4776af546b1d54def1df |
| Accession Number: |
edsdoj.3df09aa019eb4776af546b1d54def1df |
| Database: |
Directory of Open Access Journals |
