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

溶酶体膜裂变缺陷介导阿尔茨海默病营养不良神经突中轴突溶酶体的积累。

• 批准号: 10406342
• 负责人: Matthew Schrag
• 金额: $ 24.3万
• 依托单位: VANDERBILT UNIVERSITY MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-08-01 至 2024-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10406342
• 关键词: Abeta clearance; Adult; Age-Months; Aging; Alzheimer' s Disease; Alzheimer' s disease model; Alzheimer' s disease risk; Amyloid beta-Protein; Amyloidosis; Animal Model; Area; Axon; Behavioral; Biology; Blood Vessels; Brain; Caregiver Burden; Caregivers; Cell physiology; Cells; Cerebral Amyloid Angiopathy; Cerebrum; Clinical Management; Clinical Trials; Code; Cognition; Cognitive; Cognitive aging; Communities; Complex; Confocal Microscopy; Coupled; Custom; Dementia; Deposition; Deterioration; Development; Disease; Elderly; Electron Microscopy; Environment; Enzymes; Epidemic; Failure; Functional disorder; Genes; Genetic Models; Healthcare; Hela Cells; Human; Image Analysis; Immune system; Impaired cognition; Impairment; International; Investigational Therapies; Lead; Learning; Lysosomes; Mediating; Membrane; Membrane Fusion; Memory; Mentorship; Molecular; Mus; Mutate; Mutation; Nerve Degeneration; Neurites; Neurobiology; Neurons; Outcome; Pathogenesis; Pathologic; Pathology; Performance; Phenotype; Phospholipase; Phospholipase D; Phospholipases A; Plasmids; Process; Proteins; Public Health; Publishing; Regulation; Reporting; Research; Resources; Role; Running; Senile Plaques; System; Techniques; Testing; Therapeutic; Tissue Model; Training; Variant; WFS1 gene; Work; abeta deposition; aging brain; animal model development; base; behavioral phenotyping; brain tissue; career development; cerebral amyloidosis; cohort; conditional knockout; cost; density; effective therapy; enzyme activity; genetic risk factor; human tissue; improved; in vitro Assay; in vivo; link protein; lysosome membrane; mouse model; neurobehavioral; neuropathology; new therapeutic target; novel; programs; protein aggregation; protein function; proteostasis; quantitative imaging; selective expression; skills; stem; β-amyloid burden

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.

阿尔茨海默病（AD）损害了老年人的独立性，并与巨大的社会影响有关。 医疗保健和护理费用的负担。迫切需要一种有效的治疗方法来阻止这种潮流。 持续增长的痴呆症流行病最近的许多临床试验都试图降低β- 淀粉样蛋白在大脑中，通常通过激活免疫系统，以促进清除，但没有这些试验 已经成功了我们建议以不同的方式探索这个问题-我们发现神经元 β-淀粉样蛋白沉积物周围的轴突肿胀并充满异常溶酶体， 蛋白质降解酶。由于溶酶体对蛋白质稳态至关重要，我们假设， 这些异常的溶酶体会导致神经变性，如果它们的功能能够被挽救， 改善大脑功能。我们将通过关注一个新的基因PLD 3来研究这一假设，该基因已被鉴定为 增加AD风险。在我们的初步工作中，我们发现PLD 3在这些 异常溶酶体和脑PLD 3水平与β-淀粉样蛋白负荷和 认知能力的衰退。大脑PLD 3水平也与记忆力呈负相关 在小鼠模型中的表现。我们发现PLD 3在酸性环境中作为磷脂酶D发挥功能， 是溶酶体膜分裂所必需的。我们建议评估在多大程度上 在小鼠中观察到的溶酶体功能障碍存在于人体组织中， PLD 3与神经元溶酶体病理。然后，我们将确定报告给 通过转染含有PLD 3的质粒（其已被 突变以复制这些编码变体）并观察对溶酶体膜分裂和对PLD 3的影响 酶活性我们将确定是否有缺陷的神经元溶酶体分裂影响认知和AD 通过将遗传性AD的5xFAD模型与图4的条件性敲除交叉，使用免疫组织化学方法在体内观察神经病理学。 Cre选择性表达于脑神经元。图4是PIKfyve复合物的组分和图4的损失。 导致溶酶体分裂缺陷。重要的是，PIKfyve复合物还调节PLD 3加工， 改变PLD 3活性。我们将评估这只小鼠的学习和记忆能力， 溶酶体分裂影响β-淀粉样变性和营养不良性神经突。这些研究将帮助我们了解这两个 这种蛋白质在AD中的作用以及溶酶体膜融合在认知老化中的作用 的AD。平行培训计划将支持我在模型动物发育和行为方面的培训。 表型，发展先进的技能，在溶酶体神经生物学和完善的临床管理老化 所有这些都是我职业发展的关键步骤。总的来说， 机构环境、资源和跨学科导师团队， 这些专业领域将加快我获得向独立过渡所需的专业知识。