The development and application of TRUPATH biosensors to the study of GPCR pharmacology and functional selectivity

G-protein-coupled receptors (GPCRs) remain major drug targets, despite an incomplete understanding of how they signal through 16 non-visual G-protein signal transducers (collectively named the transducerome) to exert their actions. Recent efforts in GPCR drug discovery have included the design or discovery of drugs that alter the engagement of specific subsets of these pathways—a concept termed functional selectivity or “bias.” However, traditional pharmacological tools to classify and rank the pharmacological activity of test compounds have largely depended on intracellular signaling pathways downstream of receptor activation and are heavily dependent on the intracellular milieu unique to each cell type or tissue, i.e. “system bias.” Because multiple transducers can link to common pathways, the disambiguation of specific G protein pathways can be especially laborious. To overcome this, multiple teams have developed FRET and BRET tools that directly measure ligand-mediated receptor-transducer engagement, though these have been largely incomplete or incompletely characterized. This talk will describe the development, characterization, and validation of the largest set of GPCR-G protein biosensors to date, collectively dubbed TRUPATH. Specific emphasis will be placed on the versatility of these types of tools in drug-screening and receptor-characterization efforts. These biosensors are freely and openly available to academic researchers for modification and use through an MTA with the University of North Carolina at Chapel Hill or through distribution by Addgene, the non-profit plasmid repository.

Learning Objectives:

1. Describe the mechanism of G protein signalling and by what properties are they classified into their families

2. What are some key considerations when defining FRET or BRET probes?

3. In your own words, define or describe functional selectivity