Multiplexed profiling of GPCR activities by combining split TEV assays and EXT-based barcoded readouts
| Autor: | Sven P. Wichert, Sabrina Galinski, Moritz J. Rossner, Michael C. Wehr |
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| Rok vydání: | 2018 |
| Předmět: |
0301 basic medicine
Multidisciplinary lcsh:R lcsh:Medicine GPCR activity Computational biology Biology PC12 Cells Multiplexing Article Rats Receptors G-Protein-Coupled 03 medical and health sciences 030104 developmental biology Drug development Cell Line Tumor Animals Profiling (information science) lcsh:Q Biological Assay lcsh:Science Receptor Antipsychotic Agents Signal Transduction G protein-coupled receptor |
| Zdroj: | Scientific Reports Scientific Reports, Vol 8, Iss 1, Pp 1-11 (2018) |
| ISSN: | 2045-2322 |
| Popis: | G protein-coupled receptors (GPCRs) are the largest class of cell surface receptors and are implicated in the physiological regulation of many biological processes. The high diversity of GPCRs and their physiological functions make them primary targets for therapeutic drugs. For the generation of novel compounds, however, selectivity towards a given target is a critical issue in drug development as structural similarities between members of GPCR subfamilies exist. Therefore, the activities of multiple GPCRs that are both closely and distantly related to assess compound selectivity need to be tested simultaneously. Here, we present a cell-based multiplexed GPCR activity assay, termed GPCRprofiler, which uses a β-arrestin recruitment strategy and combines split TEV protein-protein interaction and EXT-based barcode technologies. This approach enables simultaneous measurements of receptor activities of multiple GPCR-ligand combinations by applying massively parallelized reporter assays. In proof-of-principle experiments covering 19 different GPCRs, both the specificity of endogenous agonists and the polypharmacological effects of two known antipsychotics on GPCR activities were demonstrated. Technically, normalization of barcode reporters across individual assays allows quantitative pharmacological assays in a parallelized manner. In summary, the GPCRprofiler technique constitutes a flexible and scalable approach, which enables simultaneous profiling of compound actions on multiple receptor activities in living cells. |
| Databáze: | OpenAIRE |
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