Elucidating the role of SORL1 as an APOE receptor in human astrocytes

阐明 SORL1 作为人星形胶质细胞中 APOE 受体的作用

• 批准号: 9983410
• 负责人: Hyo Lee
• 金额: $ 3.88万
• 依托单位: HARVARD MEDICAL SCHOOL
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-06-01 至 2022-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9983410
• 关键词: Abeta clearance; Address; Affect; Alzheimer' s Disease; Alzheimer' s disease patient; Alzheimer' s disease risk; Amyloid beta-Protein; Apolipoproteins; Astrocytes; Binding; Biochemical; Biological; Biological Assay; Biology; Biotinylation; Blood - brain barrier anatomy; Brain; CRISPR/Cas technology; Cell surface; Cells; Cerebrospinal Fluid; Characteristics; Cholesterol Homeostasis; Co-Immunoprecipitations; Collecting Cell; Complex; Confocal Microscopy; Data; Dementia; Drainage procedure; Endocytosis; Endosomes; Endothelial Cells; Enzyme-Linked Immunosorbent Assay; Family; Gene Family; Generations; Genes; Genetic; Genetic Transcription; Genetic study; Human; Individual; Inflammation; Intercellular Fluid; Knock-out; LDL-Receptor Related Protein 1; Label; Late Onset Alzheimer Disease; Lead; Ligand Binding; Ligands; Link; Lipoprotein Receptor; Lipoproteins; Low Density Lipoprotein Receptor; Lysosomes; Measures; Mediating; Meta-Analysis; Missense Mutation; Mus; Mutation; Nervous system structure; Neurodegenerative Disorders; Neurofibrillary Tangles; Neurons; Pathology; Pathway Analysis; Pathway interactions; Patients; Play; Polymorphism Analysis; Presenile Alzheimer Dementia; Property; Protein Family; Protein Isoforms; Proteins; RNA Splicing; Risk; Role; Senile Plaques; Technology; Testing; Transcript; Variant; Work; abeta accumulation; amyloid precursor protein processing; apolipoprotein E-4; base; brain cell; causal variant; cell type; experimental study; extracellular; familial Alzheimer disease; genetic risk factor; genome wide association study; human subject; induced pluripotent stem cell; loss of function; neuron loss; novel; preference; protein expression; receptor; risk variant; stem cell technology; tau Proteins; trafficking; transcriptome sequencing; uptake

Project Summary/Abstract Alzheimer’s disease (AD) is a progressive neurodegenerative disease affecting more than 30 million people worldwide. It is the most common cause of dementia, and its pathology includes extensive neuronal loss, accumulation of extracellular amyloid-beta (Aβ) plaques and intracellular neurofibrillary tangles. A number of large scale meta-analyses of genome wide association studies (GWAS) has analyzed polymorphisms in many human subjects, and identified more than 20 genes associated with AD risk, which can be classified based on their known functions: cholesterol metabolism, inflammation, and endocytosis. Our lab seeks to utilize induced pluripotent stem cells (iPSCs) from patients with familial and sporadic AD to understand the cellular mechanisms underlying AD. My proposed project will investigate the potential convergent mechanisms of APOE and SORL1 function in the brain. Both APOE and SORL1 are AD risk genes, and APOE (ε4) is the strongest genetic risk factor for AD. Not only is SORL1 a GWAS hit for late onset AD (LOAD), but recent studies identified SORL1 loss-of-function variants in familial AD patients, suggesting that SORL1 can be a potential causal gene for AD. Here, I will study the function of SORL1 as a receptor for APOE, which can address the potential convergent mechanism between these genes. I hypothesize that SORL1 will function as an APOE receptor in human astrocytes and regulate Aβ clearance. In support of this, publicly available RNA-seq data show that SORL1 transcript is highly expressed in astrocytes over other brain cell types. In addition, APOE has a known role in Aβ clearance, and SORL1 is in the APOE receptor gene family. These findings, however, do not directly examine the role of SORL1 in regulating Aβ uptake in astrocytes. Here, I will utilize iPSC-derived astrocytes with different SORL1 and APOE variants to study the ability of SORL1 to function as an APOE receptor and uptake Aβ. In Aim 1, I will generate iPSC-derived astrocytes and determine SORL1 expression, subcellular localization, and ligand binding characteristics. Then, In Aim 2, I will use iPSC-derived astrocytes generated from individuals with 1) different APOE isoforms 2) SORL1 knock out (KO), risk SNPs and missense mutations and 3) other AD risk SNPs. I then will meticulously characterize Aβ generation, uptake and clearance in these astrocytes. Our preliminary data suggest that SORL1 is highly expressed in human astrocytes, and SORL1 KO in these astrocytes results in elevated extracellular Aβ levels. Also, I have shown that iPSC-derived astrocytes reduce extracellular Aβ levels when treated with exogenous Aβ, opening up the possibility/potential for this assay to examine the Aβ clearance ability of cells from individuals with various genetic backgrounds. In summary, my proposal will utilize iPSC-derived astrocytes to examine the novel role of SORL1 as an APOE receptor in astrocytes, and investigate a potential convergent mechanism that APOE and SORL1 play in Aβ clearance.

项目总结/摘要 阿尔茨海默病（AD）是一种进行性神经退行性疾病，影响超过3000万人 国际吧它是痴呆症最常见的原因，其病理学包括广泛的神经元损失， 细胞外β淀粉样蛋白（Aβ）斑块和细胞内神经元缠结的积累。一些 全基因组关联研究（GWAS）的大规模荟萃分析已经分析了许多基因组中的多态性， 人类受试者，并确定了20多个与AD风险相关的基因，这些基因可以根据 它们的已知功能：胆固醇代谢、炎症和内吞作用。我们的实验室试图利用诱导 来自家族性和散发性AD患者的多能干细胞（iPSC），以了解细胞机制 基础AD我提出的项目将研究APOE和SORL 1的潜在会聚机制 在大脑中的功能。APOE和SORL 1都是AD的危险基因，其中APOE（ε4）的遗传危险性最大 AD的因素。SORL 1不仅是晚发性AD（LOAD）的GWAS靶点，而且最近的研究发现SORL 1 家族性AD患者中的功能缺失变异，表明SORL 1可能是AD的潜在致病基因。 在这里，我将研究SORL 1作为APOE受体的功能，它可以解决潜在的会聚性。 这些基因之间的联系。我假设SORL 1在人类中作为APOE受体发挥作用， 星形胶质细胞和调节Aβ清除。为了支持这一点，公开的RNA-seq数据显示，SORL 1 转录物在星形胶质细胞中的表达高于其他脑细胞类型。此外，已知APOE在Aβ SORL 1属于APOE受体基因家族。然而，这些发现并没有直接检查 SORL 1在调节星形胶质细胞Aβ摄取中的作用。在这里，我将利用iPSC衍生的星形胶质细胞， SORL 1和APOE变体研究SORL 1作为APOE受体和摄取Aβ的能力。在Aim中 1，我将产生iPSC衍生的星形胶质细胞，并确定SORL 1表达，亚细胞定位和配体 绑定特性然后，在目标2中，我将使用从具有1） 2）SORL 1敲除（KO）、风险SNP和错义突变和3）其他AD风险 SNPs。然后，我将仔细描述这些星形胶质细胞中Aβ的产生、摄取和清除。我们 初步数据表明，SORL 1在人类星形胶质细胞中高度表达，而SORL 1 KO在这些星形胶质细胞中表达。 星形胶质细胞导致细胞外Aβ水平升高。此外，我已经证明iPSC衍生的星形胶质细胞减少了 当用外源性Aβ处理时，细胞外Aβ水平，为该测定开辟了可能性/潜力， 检查来自不同遗传背景个体的细胞的Aβ清除能力。总之，我的 一项提案将利用iPSC衍生的星形胶质细胞来研究SORL 1作为APOE受体的新作用 研究APOE和SORL 1在Aβ中的潜在会聚机制 间隙