Developing an <i>in vivo</i> Bioluminescent Assay Platform for Determining Ligand-Induced GPCR/G-Protein Activation

<p dir="ltr">Neurodegenerative diseases are quickly becoming a leading cause of death worldwide. These diseases include Alzheimer’s disease (AD), Parkinson’s, Amyotrophic lateral sclerosis (ALS), and many others. Despite their growing prevalence, effective treatments remain elusive, with current therapies offering only limited symptomatic relief. Proteins, particularly membrane receptors, may play a central role in both the progression and potential treatment of these disorders. Among them, G-protein-coupled receptors (GPCRs) represent the largest class of druggable membrane proteins and are highly attractive targets due to their extracellular accessibility and diverse physiological roles. However, only 16–20% of the approximately 800 known GPCRs are currently targeted by approved therapeutics.</p><p dir="ltr">Orphan GPCRs (oGPCRs), GPCRs having unknown endogenous ligands, make up a significant amount of the known GPCRs. Notably, many oGPCRs are enriched in brain tissue and may hold untapped therapeutic potential for neurological diseases. Because the ligands to activate these oGPCRs remain unknown, efforts to identify natural or synthetic small molecules or peptides to activate these oGPCRs in a therapeutic manner are currently underway. </p><p dir="ltr">I developed an in vivo bioluminescent assay platform to analyze the GPCR/G-protein coupling of four oGPCRs that are predominantly expressed in brain tissue: GPR26, GPR173, GPR171, and GPR83. Elucidating their binding partners may lead to the improvement of treatments for neurological diseases. This bioluminescent assay platform will elucidate transient interactions between oGPCRs and their binding partners. </p><p dir="ltr">Using this bioluminescent assay platform, I confirmed the recent deorphanization of receptors GPR173, GPR171, and GPR83 using their identified ligands, and discovered novel G-protein coupling profiles not previously reported. The assay platform offers greater sensitivity and temporal resolution than conventional signaling assays and can be extended for high-throughput drug screening applications. </p><p dir="ltr">Together, this work provides new tools and insights into GPCR biology, and contributes to the search for novel therapeutics targeting neurodegenerative diseases.</p>