A Scalable, Multiplexed Assay for Decoding GPCR-Ligand Interactions with RNA Sequencing

G-protein coupled receptors (GPCRs) are central to how mammalian cells sense and respond to chemicals. Although 34% of FDA-approved drugs target GPCRs, the majority remain “orphaned,” that is, bonafide ligands have not been identified. Here, we introduce a platform to screen large chemical panels against multiplexed GPCR libraries using next-generation sequencing of barcoded genetic reporters in stably-engineered human cell lines. We validate this platform using mammalian olfactory receptors (ORs), a large family of GPCRs, mediate the sense of smell through activation by odorant small molecules. We mapped 39 mammalian olfactory receptors against 181 odorants and identified 79 interactions that have not been reported to our knowledge, including ligands for previously orphaned receptors. This multiplexed receptor assay allows the cost-effective mapping of large chemical libraries to receptor repertoires at scale.