A fluorescent reporter system for scalable and live detection of PYY production from enteroendocrine cells with single-event resolution

Peptide YY (PYY) comprises the secretory repertoire of enteroendocrine L-cells alongside glucagon-like peptide-1 (GLP-1), and positively modulates postprandial satiety, digestion mechanics, and regeneration of the intestinal epithelium. Whereas immortalised GLP-1-secreting human L-cell models support pre-clinical drug discovery, comparable human lines that robustly secrete PYY are lacking, hindering mechanistic studies of its release. We present a biosensor for scalable detection of PYY production and secretion from human enteroendocrine cells in vitro. Guided by in silico structural prediction, we engineer a superecliptic phluorin (SEP)-tagged PYY, SEP-PYY, that engages native hormone processing machinery and is responsive to canonical nutrient stimuli when expressed in a human enteroendocrine cell line, NCI-H716. SEP-PYY production and secretion can be measured by optimised flow cytometry and spectrofluorometric plate readouts respectively, methods with superior time- and cost-efficacy to current hormone detection methods. Leveraging the pH sensitivity of SEP, we use this reporter system in detection of single-event hormone exocytosis by total internal reflection microscopy. Finally, we demonstrate the application of our system in screening ligands of metabolite-sensing G-protein coupled receptors that drive SEP-PYY secretion, and supporting discovery of druggable pathways in metabolic disease.