Defective lysosomal membrane fission mediates axonal lysosome accumulation in dystrophic neurites in Alzheimer's disease.

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Alzheimer’s disease (AD) compromises the independence of aging adults and is associated with huge societal burden in healthcare and caregiver costs. An effective treatment is urgently needed to stem the tide of the ongoing and growing epidemic of dementia. Many recent clinical trials have attempted to reduce the level of β- amyloid in the brain, typically by activating the immune system to promote clearance, yet none of these trials has been successful. We propose to explore this problem in a different way – we discovered that neuronal axons around deposits of β-amyloid are swollen and filled with abnormal lysosomes which are deficient in protein-degrading enzymes. Because lysosomes are critical for protein homeostasis, we hypothesize that these abnormal lysosomes contribute to neurodegeneration and if their function could be rescued, it could improve brain function. We will study this hypothesis by focusing on a novel gene, PLD3, which was identified as contributing to AD risk. In our preliminary work, we discovered that PLD3 is robustly enriched on these abnormal lysosomes and brain PLD3 levels correlated inversely with both β-amyloid burden and the rate of cognitive deterioration in a human cohort. Brain PLD3 levels also correlated inversely with memory performance in a mouse model. We discovered that PLD3 functions as a phospholipase D in acidic environments and is necessary for lysosomal membrane fission. We propose to evaluate to what degree the lysosome dysfunction observed in mice is present in human tissue and to fully evaluate the association of PLD3 with neuronal lysosomal pathology. We will then determine whether the coding variants reported to confer AD-risk impact the function of PLD3 by transfecting plasmids containing PLD3 (which have been mutated to copy these coding variants) and observing the effect on lysosomal membrane fission and on PLD3 enzyme activity. We will determine whether defective neuronal lysosomal fission impacts cognition and AD neuropathology in vivo by crossing the 5xFAD model of genetic AD with a conditional knock-out of Fig4 using a Cre selectively expressed in cerebral neurons. Fig4 is a component of the PIKfyve complex and loss of Fig4 leads to defective lysosomal fission. Importantly, the PIKfyve complex also regulates PLD3 processing and alters PLD3 activity. We will evaluate learning and memory in this mouse and determine whether defective lysosomal fission impacts β-amyloidosis and dystrophic neurites. These studies will help us understand both how this protein functions in the setting of AD and the role of lysosome membrane fusion in cognitive aging and AD. The parallel training plan will support my training in model animal development and behavioral phenotyping, developing advanced skills in lyosomal neurobiology and refine clinical management of aging related diseases, all of which are critical steps for my career development. Collectively, the outstanding institutional environment, resources, and interdisciplinary mentorship team with broad and complementary areas of expertise will accelerate my acquisition of the necessary expertise to transition to independence.