Elucidating molecular mechanisms of lysosomal dysfunction underlying progranulin deficiency

阐明颗粒体蛋白前体缺乏引起的溶酶体功能障碍的分子机制

• 批准号: 10686234
• 负责人: Jessica Turner Root
• 金额: $ 2.96万
• 依托单位: EMORY UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-08-23 至 2023-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10686234
• 关键词: Affect; Alzheimer' s Disease; Birth; Brain; Cathepsins; Cell Line; Cell model; Cells; Ceramides; Cognitive; DNA Sequence Alteration; Data; Deterioration; Development; Digestion; Disease; Disease Marker; Dose; Early identification; Enzymes; Etiology; Fibroblasts; Frontotemporal Dementia; Functional disorder; GRN gene; Genetic; Glycoproteins; Health; Heterozygote; Homeostasis; Human; Hydrolase; Immune; Immunoprecipitation; Impairment; Individual; Inflammation; Inflammatory; Inflammatory Response; Injections; Learning; Length; Lipids; Liver; Lysosomes; Mass Spectrum Analysis; Mediating; Messenger RNA; Microglia; Modeling; Molecular; Mus; Mutation; Nerve Degeneration; Neurodegenerative Disorders; Neuroglia; Neuronal Ceroid-Lipofuscinosis; Neuronal Dysfunction; Neurons; Organelles; PGRN gene; Parkinson Disease; Pathogenesis; Pathogenicity; Pathologic; Pathway interactions; Patients; Peptide Hydrolases; Phagocytosis; Phenotype; Plasma; Play; Process; Proteins; Proteolysis; Proteome; Recombinant adeno-associated virus (rAAV); Recombinants; Recycling; Reporting; Risk; Role; Signal Transduction; Sphingolipids; Synapses; System; Testing; Therapeutic; Tissues; Work; aged; brain health; brain tissue; cell type; cytokine; disease phenotype; enzyme activity; experimental study; extracellular; genetic variant; granulin; in vitro Model; in vivo; induced pluripotent stem cell; lipid metabolism; loss of function; loss of function mutation; lysosomal proteins; macromolecule; migration; neuroinflammation; neuron loss; neuronal survival; neuroprotection; neurotoxic; novel; prevent; therapeutic target

PROJECT SUMMARY Progranulin (PGRN) encoded by the gene GRN is a secreted pleiotropic protein implicated in several processes including inflammation, extracellular signaling, and neuronal survival. Heterozygous, loss of function GRN mutations cause the neurodegenerative disorder frontotemporal dementia (FTD). Genetic variants in GRN that decrease circulating levels of PGRN also increase the risk of developing Alzheimer’s disease or Parkinson’s disease. These genetic discoveries demonstrate that PGRN is important for neuronal health, but it is unclear why the loss of PGRN leads to neurodegeneration. Interestingly, recently identified homozygous GRN mutations cause neuronal ceroid lipofuscinosis (CLN11). This group of lysosomal storage disorders (LSD) presents with neurodegeneration, cognitive deterioration, and lipofuscinosis in multiple tissues. The discovery that that homozygous GRN mutations cause an LSD combined with the observation that FTD-GRN patients share CLN11 pathological features, strongly suggests lysosomal dysfunction may underlie both disorders. This proposal will help elucidate the molecular role of Progranulin in the lysosome underlying mechanisms of neurodegeneration. PGRN traffics to the lysosome where it is processed into subunit granulins. It has been suggested that granulins are neurotoxic and pro-inflammatory, but recent work from the Kukar lab and others indicates that granulins play a homeostatic role in the lysosome. Data shows that granulins are stable in the lysosome compared to full length PGRN, and that granulins are decreased in human FTD-GRN cells and brain tissue. Furthermore, the complete loss of PGRN and granulins leads to accumulation of ceramides in the brain. These data suggest that granulins may be a bioactive component of lysosomal function involved in lipid degradation, yet the ability of individual granulins to ameliorate phenotypes of PGRN deficiency in vivo in unknown. Aim 1 will assess whether the expression of individual granulins is sufficient to rescue dysregulated lysosome function, and inflammation in Grn-/- mice. Progranulin has been implicated in the inflammatory response and is highly expressed in microglia, the brain’s resident immune cells. PGRN is involved in microglia activation, phagocytosis, migration, and synapse pruning. Moreover, the loss of PGRN has been shown to cause early and selective impairments in the maturation of lysosomal cathepsins, and accumulation of lipid droplets in murine microglia suggesting that microglial lysosomes may be particularly vulnerable to the loss of PGRN. Homeostatic microglial function is critical for brain health, however the impact of PGRN deficiency on human microglia is unknown. Aim 2 will assess the functional effects of PGRN deficiency on human microglia leveraging a novel patient derived iPSC line and isogenic controls developed by the Kukar lab. These experiments will be the first to directly assess the role of granulins on the molecular, and pathological, phenotypes of PGRN-deficient systems. Evaluating the bioactivity of PGRN and granulins will elucidate disease-relevant molecular pathways, and lead to the development of effective and precise therapeutics to treat diseases caused by PGRN/granulin deficiency.

项目摘要 颗粒蛋白前体（progranulin，PGRN）是由GRN基因编码的一种分泌性多效性蛋白，参与多种生物学过程 包括炎症、细胞外信号和神经元存活。杂合子，GRN功能丧失 突变导致神经退行性疾病额颞叶痴呆（FTD）。GRN中的遗传变异， 降低PGRN的循环水平也会增加患阿尔茨海默病或帕金森病的风险 疾病这些遗传学发现证明PGRN对神经元健康很重要，但目前还不清楚 为什么PGRN的缺失会导致神经退化有趣的是，最近发现的纯合GRN突变， 导致神经元蜡样质脂褐质沉积症（CLN 11）。这组溶酶体贮积症（LSD）表现为： 神经变性、认知退化和多个组织中的脂褐质沉积。发现这一点 纯合GRN突变导致LSD，并观察到FTD-GRN患者共享CLN 11 病理特征，强烈表明溶酶体功能障碍可能是这两种疾病的基础。这项建议会 有助于阐明颗粒蛋白前体在神经变性的溶酶体基础机制中的分子作用。 PGRN运输到溶酶体，在那里它被加工成亚基颗粒蛋白。有人建议 颗粒蛋白具有神经毒性和促炎性，但Kukar实验室和其他人最近的研究表明， 颗粒蛋白在溶酶体中起稳态作用。数据显示颗粒蛋白在溶酶体中是稳定的 与全长PGRN相比，颗粒蛋白在人FTD-GRN细胞和脑组织中减少。 此外，PGRN和颗粒蛋白的完全丧失导致脑中神经酰胺的积累。这些 数据表明颗粒蛋白可能是参与脂质降解的溶酶体功能的生物活性组分， 然而，单个颗粒蛋白在体内改善PGRN缺乏表型的能力尚不清楚。目标1将 评估单个颗粒蛋白的表达是否足以挽救失调的溶酶体功能，和 Grn-/-小鼠中的炎症。颗粒蛋白前体与炎症反应有关， 在小胶质细胞中表达，小胶质细胞是大脑的常驻免疫细胞。PGRN参与小胶质细胞活化，吞噬作用， 迁移和突触修剪。此外，PGRN的缺失已被证明会导致早期和选择性的 小鼠小胶质细胞中溶酶体组织蛋白酶成熟和脂滴积累的损伤 提示小胶质细胞溶酶体可能特别易受PGRN损失的影响。稳态小胶质细胞 功能对大脑健康至关重要，然而PGRN缺乏对人类小胶质细胞的影响尚不清楚。目的 2将评估PGRN缺乏对人类小胶质细胞的功能影响，利用一种新的患者来源的 Kukar实验室开发的iPSC系和同基因对照。这些实验将首次直接评估 颗粒蛋白对PGRN缺乏系统的分子和病理表型的作用。评价 PGRN和颗粒蛋白的生物活性将阐明疾病相关的分子途径，并导致 开发有效和精确的治疗剂以治疗由PGRN/颗粒蛋白缺乏引起的疾病。