Probing PAC1 receptor activation across species with an engineered sensor

Class-B1 G-protein-coupled receptors (GPCRs) are an important family of clinically relevant drug targets that remain difficult to investigate via high-throughput screening and in animal models. Here, we engineered PAClight1 P78A , a novel genetically encoded sensor based on a class-B1 GPCR (the human PAC1 receptor, hmPAC1R) endowed with high dynamic range (Δ F / F 0 = 1100%), excellent ligand selectivity, and rapid activation kinetics ( τ ON = 1.15 s). To showcase the utility of this tool for in vitro applications, we thoroughly characterized and compared its expression, brightness and performance between PAClight1 P78A -transfected and stably expressing cells. Demonstrating its use in animal models, we show robust expression and fluorescence responses upon exogenous ligand application ex vivo and in vivo in mice, as well as in living zebrafish larvae. Thus, the new GPCR-based sensor can be used for a wide range of applications across the life sciences empowering both basic research and drug development efforts.

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Publication Details

Journal
eLife
Published
2024-08-15
DOI
https://doi.org/10.7554/elife.96496.3
Primary Topic
Receptor Mechanisms and Signaling
Type
article
Field-Weighted Citation Impact
0.00

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article

Probing PAC1 receptor activation across species with an engineered sensor

Gil Levkowitz, Preethi Rajamannar, Kevin Assoumou, Andrea Gresch et al.
eLife
Receptor Mechanisms and Signaling
article

Probing PAC1 receptor activation across species with an engineered sensor

Gil Levkowitz, Preethi Rajamannar, Kevin Assoumou, Andrea Gresch, Elyse T. Williams, Dietmar Benke, Musadiq A. Bhat, Sarah Melzer, Reto B. Cola, Nina Hartrampf, Tommaso Patriarchi, Miriam Stoeber, Salome N Niethammer, Patrick Hauck
article en

Abstract

Class-B1 G-protein-coupled receptors (GPCRs) are an important family of clinically relevant drug targets that remain difficult to investigate via high-throughput screening and in animal models. Here, we engineered PAClight1 P78A , a novel genetically encoded sensor based on a class-B1 GPCR (the human PAC1 receptor, hmPAC1R) endowed with high dynamic range (Δ F / F 0 = 1100%), excellent ligand selectivity, and rapid activation kinetics ( τ ON = 1.15 s). To showcase the utility of this tool for in vitro applications, we thoroughly characterized and compared its expression, brightness and performance between PAClight1 P78A -transfected and stably expressing cells. Demonstrating its use in animal models, we show robust expression and fluorescence responses upon exogenous ligand application ex vivo and in vivo in mice, as well as in living zebrafish larvae. Thus, the new GPCR-based sensor can be used for a wide range of applications across the life sciences empowering both basic research and drug development efforts.

eLifeVol. 13
University of Geneva (CH), University of Zurich (CH), ETH Zurich (CH), Institute of Pharmacology (CZ), Weizmann Institute of Science (IL), Medical University of Vienna (AT)
Schweizerischer Nationalfonds zur Förderung der Wissenschaftlichen Forschung, Vienna Science and Technology Fund, Israel Science Foundation, Olga Mayenfisch Stiftung, Benoziyo Endowment Fund for the Advancement of Science
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Receptor Mechanisms and Signaling
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