Investigating the role of an EIF2B3 variant as an Alzheimer's disease risk modifier

研究 EIF2B3 变体作为阿尔茨海默病风险调节剂的作用

基本信息

批准号：
10680062
负责人：
Francesca Garretti
金额：
$ 6.91万
依托单位：
ICAHN SCHOOL OF MEDICINE AT MOUNT SINAI
依托单位国家：
美国
项目类别：
财政年份：
2023
资助国家：
美国
起止时间：
2023-03-23 至 2026-03-22
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10680062
关键词：
Affect Age Age Years Alleles Alzheimer&apos s Disease Alzheimer&apos s disease patient Alzheimer&apos s disease risk Amino Acid Substitution Amyloid deposition Apolipoprotein E Bacteria Biological Assay Brain CRISPR/Cas technology Cell physiology Cells Cellular Stress Cholesterol Clinical Cognitive Complex Critical Thinking Deposition Disease EIF2B2 gene Exhibits Fellowship Fluorescence Genes Genetic study Genotype Goals Guanine Guanine Nucleotide Exchange Factors Hematopoietic stem cells High Pressure Liquid Chromatography Homeostasis Human Human Genetics Immune Immunocompromised Host Immunohistochemistry Impaired cognition Impairment In Vitro Individual Inflammation Inflammatory Injections Knowledge Measures Mediating Mentorship Messenger RNA Methods Microglia Morphology Mus Mutation Myelin Nerve Degeneration Outcome Pathway interactions Patients Phagocytes Pharmaceutical Preparations Phenotype Phosphorylation Production Productivity Proliferating Protein Biosynthesis Protein Isoforms Proteins Public Health Research Personnel Risk Role Scientist Senile Plaques Single Nucleotide Polymorphism Stress Structural Chemistry Structure Technical Expertise Techniques Testing Thapsigargin Therapeutic Training Translations Transplantation Variant Work Xenograft procedure activating transcription factor 4 biological adaptation to stress career collaborative environment cytokine disorder risk efficacy testing enzyme activity genetic risk factor genetic variant genome wide association study humanized mouse improved in vitro activity in vivo induced pluripotent stem cell inhibitor inhibitor therapy innovation live cell imaging medical schools mouse model mutant novel novel therapeutics precursor cell protein purification response risk variant single nucleus RNA-sequencing skills stressor transcription factor transcriptomics

项目摘要

Project Summary Human genetic studies provide a powerful approach to identify genes and pathways associated with Alzheimer's disease (AD). Apolipoprotein E isoform 4 (APOE4) is the strongest genetic risk factor for sporadic Alzheimer's disease. Despite the strong effect of APOE4 on AD risk, a wide range of clinical outcomes exist within APOE4 carriers: some can develop cognitive impairment as early as 30 years of age while other carriers remain cognitively normal beyond 100 years. The large range in clinical manifestation amongst APOE4 carriers suggests that there are genetic variants that modulate APOE-associated risk. To investigate this hypothesis, we conducted stratified genome-wide association studies of APOE4 carriers at the extremes of age at onset distribution to identify variants that modify risk. We identified a single variant in EIF2B3 encoding the amino acid substitution S404A as a candidate risk modifier. eIF2B3 is a subunit of eIF2B, a guanine exchange factor (GEF) for eIF2 that is involved in translational control and the integrated stress response. Our lab has recently shown that APOE4 induces a constitutively upregulated ISR and aberrant protein translation in murine and human iPSC-derived microglia, demonstrating that APOE4 alone can lead to increased levels of cellular stress1,2. Furthermore, EIF2B2, another eIF2B subunit, is an AD risk gene identified in AD GWAS3. The overall goal of this proposal is to directly test the hypothesis that EIF2B3 variants modulate APOE4-associated risk by dysregulating the ISR and thus alter microglial cell function in the context of disease. In Aim 1, I will combine cell-free in vitro fluorescent GEF assays and structural chemistry to determine the effects of EIF2B3S404A directly on eIF2B structural integrity and enzymatic activity, in the absence of cellular stress. Aim 2 will determine the impact of EIF2B3S404A on APOE4 microglia cell function in vitro using human induced pluripotent stem cell (iPSCs)- derived microglia and live-cell imaging in the context of disease-related stressors. Lastly, in Aim 3, I will employ novel xenotransplantation methods involving direct injection of human microglia precursor cells into the mouse brain to evaluate the effect of EIF2B3S404A on APOE4 microglia cell function in vivo and in the context of disease. Together, the results here will confirm or refute the hypothesis that EIF2B3 variants modulate APOE4-associated risk by altering microglial cell function. Throughout the project, I will test the efficacy of an ISR inhibitor (ISRIB) drug at alleviating EIF2B3S404A -induced effects on EIF2B activity in vitro and in microglia in vitro and in vivo. The work here has the potential to greatly impact public health through understanding the way in which variants affect immune cell function and modify disease risk and by potentially identifying a novel drug for the treatment of AD. The fellowship training plan proposed here will provide me with the technical skills, scientific knowledge, critical thinking skills gained from Dr. Alison Goate's mentorship and the collaborative environment at the Icahn School of Medicine at Mount Sinai at large, equipping me with the necessary skills for a successful transition to a career as an independent scientist.

项目概要人类遗传学研究提供了一种强大的方法来识别与阿尔茨海默病相关的基因和途径疾病（AD）。载脂蛋白 E 同工型 4 (APOE4) 是散发性阿尔茨海默病最强的遗传风险因素疾病。尽管 APOE4 对 AD 风险有很强的影响，但 APOE4 存在广泛的临床结果携带者：有些携带者早在 30 岁时就可能出现认知障碍，而其他携带者则仍然存在 100 年后认知能力正常。 APOE4 携带者的临床表现差异很大表明存在调节 APOE 相关风险的遗传变异。为了研究这个假设，我们进行了 APOE4携带者在发病年龄极端分布的分层全基因组关联研究识别改变风险的变体。我们在 EIF2B3 中鉴定出编码氨基酸取代的单个变体 S404A 作为候选风险调节剂。 eIF2B3 是 eIF2B 的一个亚基，eIF2B 是 eIF2 的鸟嘌呤交换因子 (GEF)，参与翻译控制和综合应激反应。我们的实验室最近表明 APOE4 在小鼠和人类 iPSC 衍生中诱导组成性上调 ISR 和异常蛋白翻译小胶质细胞，证明 APOE4 单独可以导致细胞应激水平增加1,2。此外， EIF2B2 是另一个 eIF2B 亚基，是 AD GWAS3 中鉴定的 AD 风险基因。该提案的总体目标是直接检验 EIF2B3 变体通过 ISR 失调调节 APOE4 相关风险的假设从而改变疾病背景下的小胶质细胞功能。在目标 1 中，我将结合无细胞体外荧光 GEF 检测和结构化学可确定 EIF2B3S404A 直接对 eIF2B 结构完整性的影响和酶活性，在没有细胞应激的情况下。目标 2 将确定 EIF2B3S404A 对使用人诱导多能干细胞 (iPSC) 衍生的小胶质细胞和 APOE4 小胶质细胞在体外发挥功能疾病相关压力源背景下的活细胞成像。最后，在目标 3 中，我将采用小说异种移植方法涉及将人类小胶质细胞前体细胞直接注射到小鼠大脑中评估 EIF2B3S404A 在体内和疾病背景下对 APOE4 小胶质细胞功能的影响。总之，这里的结果将证实或反驳 EIF2B3 变体调节 APOE4 相关的假设通过改变小胶质细胞功能来风险。在整个项目中，我将测试 ISR 抑制剂 (ISRIB) 的功效减轻 EIF2B3S404A 诱导的体外和小胶质细胞体外和体内 EIF2B 活性影响的药物。这通过了解变异影响的方式，这里的工作有可能极大地影响公共健康免疫细胞功能和改变疾病风险，并有可能确定一种治疗 AD 的新药。这里提出的进修培训计划将为我提供技术技能、科学知识、关键能力从艾莉森·戈特博士的指导和伊坎学院的协作环境中获得的思维技能西奈山医学博士，为我提供了成功过渡到职业生涯所需的技能作为一名独立科学家。