G protein-coupled receptors (GPCRs) are central mediators of mammalian cellsÂ’ ability to sense and respond to their environment. The 813 human GPCRs are the largest class of membrane receptors, are central mediators of cell physiology, and are the target of ~34% of all U.S. Food and Drug Administration (FDA)-approved drugs and ~60% of prescriptions. Here we will leverage recent advances in DNA synthesis, genome editing, next-generation DNA sequencing, and informatics develop a platform to build thousands of individual mutations to GPCRs and experimentally characterize their effects in a novel assay that can be done in a simple pooled format in human cell lines. We will develop methodologies to characterize how these mutational libraries functionally signal through the two major pathways that GPCRs signal through, the cyclic AMP and calcium signaling pathways. The profiles will give us the ability to better understand the functional consequences of genetic variation in GPCRs and how they might be impacting diseases and drug responses. In addition, analysis of these mutational profiles will give us insights on how to target these receptors for making new drugs. The technologies developed here should be broadly and directly applicable to the vast majority of GPCRs, as well as other important classes of drug targets where function can be assessed by transcriptional reporters in human cell lines.
Public Health Relevance Statement: Project narrative: Functional Genomics of G Protein-Coupled Receptors Determining the functional and molecular consequences of genetic variation amongst people will be critical to understand how such variation impacts human health, disease, and treatment. Here we are building very high-throughput methods that will allow us to functionally characterize almost all genetic variants in the largest class of druggable receptors called G Protein-Coupled Receptors. These methods are generalizable to many other important classes of proteins that affect human physiology to provide a scalable way for us to interpret the effect of genetic variation on human physiology, help us develop better drugs, and know how to better target them to the right populations.
Project Terms: Affect; Agonist; alpha-Melanocyte stimulating hormone; Amino Acids; Amplifiers; Bar Codes; base; Binding; Biological Assay; Blindness; burden of illness; Calcium; Calcium Signaling; Calcium-Sensing Receptors; Cell Line; Cell physiology; Cells; Chemicals; Complex; Cyclic AMP; Cyclic AMP Receptors; design; Disease; DNA biosynthesis; DNA sequencing; Drug Targeting; Engineering; Environment; Epitopes; functional genomics; G Protein-Coupled Receptor Signaling; G-Protein-Coupled Receptors; gain of function; gene function; Genetic; Genetic Transcription; genetic variant; Genetic Variation; genome editing; Genome engineering; Health; Hormones; Human; Human Cell Line; improved; Individual; Informatics; innovation; insight; Libraries; loss of function; Mammalian Cell; Measures; Mediator of activation protein; Melanocortin 4 Receptor; Membrane; Methodology; Methods; Missense Mutation; Molecular; mutant; Mutation; Mutation Analysis; Natural Product Drug; next generation; next generation sequencing; Nonsense Mutation; novel; novel therapeutics; Obesity; Odors; Oligonucleotides; Output; Pathway interactions; Pharmaceutical Preparations; Pharmacogenomics; Phase; Physiological; Physiology; Play; Population; protective effect; Proteins; Proxy; receptor; receptor expression; receptor function; Receptor Signaling; Reporter; response; Role; Running; Signal Pathway; Signal Transduction; Stimulus; System; targeted treatment; Technology; technology development; Therapeutic Intervention; United States Food and Drug Administration; Variant
