TRH receptor mobility in the plasma membrane is strongly affected by agonist binding and by interaction with some cognate signaling proteins
Language English Country Great Britain, England Media print-electronic
Document type Journal Article
- Keywords
- FRAP, Gq/11 protein, TRH receptor, lateral mobility, phospholipase Cβ1, β-arrestin,
- MeSH
- beta-Arrestins chemistry genetics MeSH
- Cell Membrane drug effects MeSH
- Fluorescence Recovery After Photobleaching MeSH
- HEK293 Cells MeSH
- Thyrotropin-Releasing Hormone chemistry metabolism MeSH
- G-Protein-Coupled Receptor Kinase 2 chemistry MeSH
- Humans MeSH
- Ligands MeSH
- Midazolam pharmacology MeSH
- GTP-Binding Protein alpha Subunits, Gi-Go chemistry genetics MeSH
- Receptors, Thyrotropin-Releasing Hormone agonists antagonists & inhibitors chemistry genetics MeSH
- Signal Transduction drug effects genetics MeSH
- Protein Binding drug effects MeSH
- Binding Sites MeSH
- Check Tag
- Humans MeSH
- Publication type
- Journal Article MeSH
- Names of Substances
- beta-Arrestins MeSH
- Thyrotropin-Releasing Hormone MeSH
- G-Protein-Coupled Receptor Kinase 2 MeSH
- Ligands MeSH
- Midazolam MeSH
- GTP-Binding Protein alpha Subunits, Gi-Go MeSH
- Receptors, Thyrotropin-Releasing Hormone MeSH
OBJECTIVES: Extensive research has been dedicated to elucidating the mechanisms of signal transduction through different G protein-coupled receptors (GPCRs). However, relatively little is known about the regulation of receptor movement within the cell membrane upon ligand binding. In this study we focused our attention on the thyrotropin-releasing hormone (TRH) receptor that typically couples to Gq/11 proteins. METHODS: We monitored receptor diffusion in the plasma membrane of HEK293 cells stably expressing yellow fluorescent protein (YFP)-tagged TRH receptor (TRHR-YFP) by fluorescence recovery after photobleaching (FRAP). RESULTS: FRAP analysis indicated that the lateral movement of the TRH receptor was markedly reduced upon TRH binding as the value of its diffusion coefficient fell down by 55%. This effect was prevented by the addition of the TRH receptor antagonist midazolam. We also found that siRNA-mediated knockdown of Gq/11α, Gβ, β-arrestin2 and phospholipase Cβ1, but not of Giα1, β-arrestin1 or G protein-coupled receptor kinase 2, resulted in a significant decrease in the rate of TRHR-YFP diffusion, indicating the involvement of the former proteins in the regulation of TRH receptor behavior. The observed partial reduction of the TRHR-YFP mobile fraction caused by down-regulation of Giα1 and β-arrestin1 suggests that these proteins may also play distinct roles in THR receptor-mediated signaling. CONCLUSION: These results demonstrate for the first time that not only agonist binding but also abundance of some signaling proteins may strongly affect TRH receptor dynamics in the plasma membrane.
References provided by Crossref.org
Biochemical and physiological insights into TRH receptor-mediated signaling
