PROJECT SUMMARY Tissue stem cell lineages maintain organ function and respond to dietary and physiological factors, and they also experience intrinsic metabolic shifts as cells differentiate Metabolic/physiological alterations are also linked to a number of diseases, including obesity and cancer. The mechanisms associated with intrinsic metabolic shifts in stem cell lineages and how physiological factors affect them remain largely underexplored. The Drosophila melanogaster ovary is ideal for the study of metabolic changes and their systemic control. Oogenesis is an energy/nutrient-intensive process that precisely links oocyte development through the germline stem cell (GSC) lineage with accumulation of lipids, carbohydrates, and other macromolecules. Developmentally-controlled metabolic changes occur along differentiation of the GSC lineage. Over the past ~18 years, our work has shed light on a multi-organ network that tightly coordinates oogenesis with whole- body physiology. GSCs and their progeny grow and divide faster on nutrient-rich rather than poor diets, and brain insulin-like peptides, Target of Rapamycin, AMP-dependent kinase, the steroid hormone ecdysone, and other factors mediate this response. Other organs also support the nutritional demands of oogenesis. For example, adipocyte lipophorin-mediated transport of lipids is crucial for oocyte yolk uptake, and several other adipocyte metabolic pathways have specific effects in oogenesis. The coordination of hormones, nutrients, metabolism, and highly regulated transitions in the GSC lineage thus integrates information from the diet and other organs. It remains unclear, however, how diet-dependent pathways affect cellular metabolism as cells differentiate along stem cell lineages, and how metabolic disorders (e.g. obesity) alter this complex process. Cellular metabolism is closely tied to nutrient fuel availability and utilization. Major cellular fuels include sugars, fatty acids, amino acids, and ketone bodies, and their availability varies depending on the overall physiological and metabolic state of the organism. Over the next 5 years, we will focus on two major questions in the Drosophila model: 1) How does fuel preference shift as GSC daughters develop through various stages of differentiation and in response to diet-dependent physiological input? 2) How does obesity impact the development of the GSC lineage, its fuel preference, and response to physiological signals? These projects will provide fundamental new knowledge to significantly advance our understanding of the integration between metabolism and physiology in the control of stem cell lineages in vivo, with the potential to inform future research additional stem cell systems and how their metabolic deregulation is tied to diseased states.