Deciphering and disabling lysosome-dependent mechanisms of neurodegeneration - The survival of neuronal cells critically depends on the correct function of their lysosomes, catabolic organelles that play key roles in disposing damaged and harmful cellular components. Niemann-Pick type C (NPC) is a neurodegenerative and metabolic disease triggered by mutations of the NPC1 gene, which encodes a lysosomal membrane protein involved in the trafficking and partitioning of cholesterol. In cells lacking NPC1, cholesterol accumulates aberrantly inside the lysosome, triggering a cascade of downstream events that impact mitochondria, ultimately leading to cellular dysfunction and death. Understanding how the molecular details of the primary lysosomal dysfunction, and the downstream processes that impact mitochondrial homeostasis and overall cell metabolism is key to the design of more effective and targeted strategies to restore neuronal cell homeostasis in NPC. Recently, the PI (RZ) and co-Investigator (PO) laboratories have made foundational discoveries that deepened the current understanding of NPC pathogenesis. In particular, through organelle immunoisolation and profiling we uncovered a profound impairment in lysosomal proteolysis, hydrolase content and membrane stability, coupled with defective delivery of damaged mitochondria to the lysosome during autophagy. Additionally, through initial studies in iPSC-derived patient-specific and NPC1-deleted neurons, we uncovered a direct connection between alterations of lysosomal function, mitochondrial metabolism and neuronal failure. Finally, we established that mTORC1, a nutrient-sensing pathway based at the lysosome, becomes dysregulated in NPC, and that its pharmacological manipulation corrects multiple organelle defects. Combined, these findings provide a rich and detailed understanding of molecular aspects of NPC pathology. Specifically, they lead us to hypothesize that the lysosomal cholesterol-mTORC1 pathway we discovered drives loss of neuronal cell homeostasis and may be a therapeutic target in NPC. In order to effectively target the cholesterol- mTORC1 axis for neuronal resilience and NPC therapy, we propose to i) mechanistically and structurally elucidate the molecular mechanisms connecting cholesterol levels to mTORC1 regulation, ii) determine how selectively disabling the cholesterol-mTORC1 sensing pathway impacts mitochondrial metabolism and neuronal survival in NPC disease models iii) discover new pathways that control organelle function and cellular metabolism, and evaluate their role as candidate NPC disease modifiers. To accomplish these goals, we will combine biochemical, cellular and unbiased approaches, including functional genomics and metabolomics in both neuronal and non-neuronal models of NPC disease. We will rigorously prioritize and validate the most promising targets, and elucidate their role within the framework of organelle function and neuronal cell homeostasis. This proposal will deepen our understanding of NPC drivers and disease modifiers, with relevance to neurodegenerative conditions linked to cholesterol imbalance, such as Alzheimer Disease.
