Bibliographic Details
| Title: |
Conformation- and activation-based BRET sensors differentially report on GPCR–G protein coupling. |
| Authors: |
Wright, Shane C., Avet, Charlotte, Gaitonde, Supriya A., Muneta-Arrate, Itziar, Le Gouill, Christian, Hogue, Mireille, Breton, Billy, Koutsilieri, Stefania, Alarcia, Rebeca Diez, Héroux, Madeleine, Lauschke, Volker M., Bouvier, Michel |
| Source: |
Science Signaling; 6/18/2024, Vol. 17 Issue 841, p1-8, 8p |
| Subject Terms: |
G protein coupled receptors, FLUORESCENCE resonance energy transfer, MUSCARINIC receptors, G proteins, CELL communication, BIOLUMINESCENCE |
| Abstract: |
G protein–coupled receptors (GPCRs) regulate cellular signaling processes by coupling to diverse combinations of heterotrimeric G proteins composed of Gα, Gβ, and Gγ subunits. Biosensors based on bioluminescence resonance energy transfer (BRET) have advanced our understanding of GPCR functional selectivity. Some BRET biosensors monitor ligand-induced conformational changes in the receptor or G proteins, whereas others monitor the recruitment of downstream effectors to sites of G protein activation. Here, we compared the ability of conformation-and activation-based BRET biosensors to assess the coupling of various class A and B GPCRs to specific Gα proteins in cultured cells. These GPCRs included serotonin 5-HT2A and 5-HT7 receptors, the GLP-1 receptor (GLP-1R), and the M3 muscarinic receptor. We observed different signaling profiles between the two types of sensors, highlighting how data interpretation could be affected by the nature of the biosensor. We also found that the identity of the Gβγ subunits used in the assay could differentially influence the selectivity of a receptor toward Gα subtypes, emphasizing the importance of the receptor-Gβγ pairing in determining Gα coupling specificity. Last, the addition of epitope tags to the receptor could affect stoichiometry and coupling selectivity and yield artifactual findings. These results highlight the need for careful sensor selection and experimental design when probing GPCR–G protein coupling. Editor's summary: Bioluminescence resonance energy transfer (BRET)–based biosensors are commonly used to determine the specific G proteins that are activated by a particular G protein–coupled receptor (GPCR). Wright et al. found that the type of biosensor and other aspects of the experimental design, such as whether the receptor was tagged and the specific Gβ and Gγ subunits expressed in the cells, influenced whether the assay detected GPCR coupling to a particular Gα subunit. These findings have the potential to improve the design of BRET-based assays to accurately determine GPCR functional selectivity. —Annalisa M. VanHook [ABSTRACT FROM AUTHOR] |
|
Copyright of Science Signaling is the property of American Association for the Advancement of Science and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.) |
| Database: |
Complementary Index |
