Role of G protein‐coupled receptor kinases (GRKs) in β2‐adrenoceptor‐mediated glucose uptake

Ham, Seungmin; Mukaida, Saori; Sato, Masaaki; Keov, Peter; Bengtsson, Tore; Furness, Sebastian; Holliday, Nicholas D.; Evans, Bronwyn A.; Summers, Roger J.; Hutchinson, Dana S.

Abstract

Truncation of the C‐terminal tail of the β2‐AR, transfection of βARKct or over‐expression of a kinase‐dead GRK mutant reduces isoprenaline‐stimulated glucose uptake, indicating that GRK is important for this response. We explored whether phosphorylation of the β2‐AR by GRK2 has a role in glucose uptake or if this response is related to the role of GRK2 as a scaffolding protein. CHO‐GLUT4myc cells expressing wild‐type and mutant β2‐ARs were generated and receptor affinity for [3H]‐CGP12177A and density of binding sites determined together with the affinity of isoprenaline and BRL37344. Following receptor activation by β2‐AR agonists, cAMP accumulation, GLUT4 translocation, [3H]‐2‐deoxyglucose uptake, and β2‐AR internalization were measured. Bioluminescence resonance energy transfer was used to investigate interactions between β2‐AR and β‐arrestin2 or between β2‐AR and GRK2. Glucose uptake after siRNA knockdown or GRK inhibitors was measured in response to β2‐AR agonists. BRL37344 was a poor partial agonist for cAMP generation but displayed similar potency and efficacy to isoprenaline for glucose uptake and GLUT4 translocation. These responses to β2‐AR agonists occurred in CHO‐GLUT4myc cells expressing β2‐ARs lacking GRK or GRK/PKA phosphorylation sites as well as in cells expressing the wild‐type β2‐AR. However, β2‐ARs lacking phosphorylation sites failed to recruit β‐arrestin2 and did not internalize. GRK2 knock‐down or GRK2 inhibitors decreased isoprenaline‐stimulated glucose uptake in rat L6 skeletal muscle cells. Thus, GRK phosphorylation of the β2‐AR is not associated with isoprenaline‐ or BRL37344‐stimulated glucose uptake. However, GRKs acting as scaffold proteins are important for glucose uptake as GRK2 knock‐down or GRK2 inhibition reduces isoprenaline‐stimulated glucose uptake.

Citation

Ham, S., Mukaida, S., Sato, M., Keov, P., Bengtsson, T., Furness, S., Holliday, N. D., Evans, B. A., Summers, R. J., & Hutchinson, D. S. (2024). Role of G protein‐coupled receptor kinases (GRKs) in β2‐adrenoceptor‐mediated glucose uptake. Pharmacology Research and Perspectives, 12(1), Article e1176. https://doi.org/10.1002/prp2.1176

• Journal Article Type: Article
• Acceptance Date: Jan 22, 2024
• Online Publication Date: Feb 8, 2024
• Publication Date: 2024-02
• Deposit Date: Feb 11, 2024
• Publicly Available Date: Feb 12, 2024
• Journal: Pharmacology Research and Perspectives
• Print ISSN: 2052-1707
• Electronic ISSN: 2052-1707
• Publisher: Wiley Open Access
• Peer Reviewed: Peer Reviewed
• Volume: 12
• Issue: 1
• Article Number: e1176
• DOI: https://doi.org/10.1002/prp2.1176
• Keywords: β2 adrenoceptor, glucose uptake, GRK2
• Public URL: https://nottingham-repository.worktribe.com/output/31160032
• Publisher URL: https://bpspubs.onlinelibrary.wiley.com/doi/10.1002/prp2.1176