Dynamic mass redistribution reveals diverging importance of PDZ-ligands for G protein-coupled receptor pharmacodynamics
| Autor: | Dorathy-Ann Harris, Nathan D. Camp, Nephi Stella, Jennifer L. Wacker-Mhyre, Kyung Soon Lee, Alejandro Wolf-Yadlin, Timothy S. Kountz, Ji Min Park, Allison E. Cherry, Aaron Stewart, Marianne Estrada, Chris Hague |
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| Rok vydání: | 2015 |
| Předmět: |
0301 basic medicine
Pharmacology LPAR2 P2RY1 PDZ domain PDZ Domains Biology Ligands Article Receptors G-Protein-Coupled 03 medical and health sciences 030104 developmental biology HEK293 Cells Membrane protein Drug Discovery Biophysics Humans Protein Interaction Domains and Motifs Receptor Cellular localization S1PR2 G protein-coupled receptor Signal Transduction |
| Zdroj: | Pharmacological research. 105 |
| ISSN: | 1096-1186 |
| Popis: | G protein-coupled receptors (GPCRs) are essential membrane proteins that facilitate cell-to-cell communication and co-ordinate physiological processes. At least 30 human GPCRs contain a Type I PSD-95/DLG/Zo-1 (PDZ) ligand in their distal C-terminal domain; this four amino acid motif of X-[S/T]-X-[φ] sequence facilitates interactions with PDZ domain-containing proteins. Because PDZ protein interactions have profound effects on GPCR ligand pharmacology, cellular localization, signal-transduction effector coupling and duration of activity, we analyzed the importance of Type I PDZ ligands for the function of 23 full-length and PDZ-ligand truncated (ΔPDZ) human GPCRs in cultured human cells. SNAP-epitope tag polyacrylamide gel electrophoresis revealed most Type I PDZ GPCRs exist as both monomers and multimers; removal of the PDZ ligand played minimal role in multimer formation. Additionally, SNAP-cell surface staining indicated removal of the PDZ ligand had minimal effects on plasma membrane localization for most GPCRs examined. Label-free dynamic mass redistribution functional responses, however, revealed diverging effects of the PDZ ligand. While no clear trend was observed across all GPCRs tested or even within receptor families, a subset of GPCRs displayed diminished agonist efficacy in the absence of a PDZ ligand (i.e. HT2RB, ADRB1), whereas others demonstrated enhanced agonist efficacies (i.e. LPAR2, SSTR5). These results demonstrate the utility of label-free functional assays to tease apart the contributions of conserved protein interaction domains for GPCR signal-transduction coupling in cultured cells. |
| Databáze: | OpenAIRE |
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