Ayala-Navarro, Carlos I2017-05-022017-05-022016-07https://hdl.handle.net/11299/187888University of Minnesota PhD dissertation. July 2016. Major: Molecular, Cellular, Developmental Biology and Genetics. Advisor: Dr. Thomas P. Neufeld. 1 computer file (PDF): v, 139 pages.Autophagy is a conserved lysosomal dependent pathway employed by cells during stress conditions as an alternative source of nutrients to maintain cellular homeostasis and promote survival. The pathway is negatively regulated by the mechanistic target of rapamycin (mTOR) and induced by depletion of nutrients. Over the last decade input in the form of vesicular traffic from an array of cellular organelles (e.g. Golgi, ER, endocytic pathway and mitochondria) has been shown to be required for the delivery of proteins, enzymes and lipids during progression of the autophagic pathway. However, how these organelles switch from their constitutive roles to supply the autophagic pathway with proteins and lipids upon induction is not fully understood. In addition the extent to which these cellular organelles modulate autophagosomal growth and mTOR-Insulin signaling remains incompletely understood. The main goal of this thesis was to uncover novel traffic regulators of the Rab GTPase family required for starvation-induced autophagy in Drosophila fat body cells and evaluate their role in mTOR signaling regulation. To this end we carried a reverse screen using RNAi to knockdown 30 of the 33 Drosophila Rab GTPases. We show Rab 2, 7 and 14 GTPases are required for the induction and growth of autophagosomes and autolysosomal function. Rab5 is required for autophagic vesicle induction and growth, endocytosis and lysosomal maturation. Lastly, that Rab6 is required for the sorting of lysosomal hydrolases, autolysosome turnover and the regulation of mTOR signaling via regulation of the insulin receptor localization in fat body cells. Altogether we uncovered novel regulators in the vesicular traffic regulator Rab GTPase family required for autophagy and mTOR-Insulin signaling regulation in Drosophila.enCellular biologyDevelopmental biologyGeneticsAutophagyInsulin signalingLysosomeNutrientsRab GTPaseTrafficThesis or Dissertation