Probing PAC1 receptor activation across species with an engineered sensor

Area of Science:

• Biochemistry
• Molecular Biology
• Pharmacology

Background:

• Class-B1 G-protein-coupled receptors (GPCRs) are crucial drug targets but challenging for high-throughput screening and animal studies.
• Investigating GPCRs requires advanced tools for in vitro and in vivo applications.

Purpose of the Study:

• To engineer a novel, genetically encoded sensor based on the human PAC1 receptor (hmPAC1R) for Class-B1 GPCRs.
• To characterize the sensor's performance for in vitro and in vivo applications, aiding drug development and basic research.

Main Methods:

• Engineered PAClight1P78A, a genetically encoded sensor derived from hmPAC1R.
• Characterized sensor expression, brightness, and performance in transfected and stably expressing cells.
• Demonstrated sensor utility in ex vivo and in vivo mouse models and in living zebrafish larvae.

Main Results:

• PAClight1P78A exhibits a high dynamic range (1100%), excellent ligand selectivity, and rapid activation kinetics (1.15 s).
• Robust sensor expression and fluorescence responses were observed in various animal models (mice and zebrafish).
• The sensor proved effective for both in vitro and in vivo applications.

Conclusions:

• PAClight1P78A is a versatile, high-performance sensor for Class-B1 GPCRs.
• This novel tool facilitates research and drug development targeting GPCRs.
• The sensor empowers a broad range of life science applications.