Modeling Alzheimer's disease genetic variants in hiPSC

在 hiPSC 中模拟阿尔茨海默病遗传变异

• 批准号: 10594510
• 负责人: Jubao Duan
• 金额: $ 62.98万
• 依托单位: NORTHSHORE UNIVERSITY HEALTHSYSTEM
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-05-01 至 2025-02-28
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10594510
• 关键词: ATAC-seq; Address; Affect; Alleles; Alzheimer' s Disease; Alzheimer' s disease model; Alzheimer' s disease risk; Astrocytes; Biochemical; Bioinformatics; Biological Assay; Biology; Brain; CRISPR interference; CRISPR/Cas technology; Cell Line; Cells; Cellular biology; Chromatin; Clustered Regularly Interspaced Short Palindromic Repeats; Coculture Techniques; Complement; Data; Data Set; Dementia; Disease; Electrophysiology (science); Exhibits; Gene Expression; Genes; Genetic Diseases; Genetic Transcription; Genome; Genomics; Glutamates; Heterozygote; Hi-C; Human; Human Engineering; Induced pluripotent stem cell derived neurons; Investigation; Knowledge; Linkage Disequilibrium; Maps; Measurement; Microglia; Molecular; Morphology; Neurodegenerative Disorders; Neuroglia; Neurons; Pathogenesis; Pathway interactions; Phenotype; Proxy; Regulation; Reproducibility; Risk; Sampling; Schizophrenia; Single Nucleotide Polymorphism; Site; System; Testing; Transcript; Transposase; Untranslated RNA; Variant; aging population; bioinformatics tool; causal variant; cell type; clinical translation; cost; effective intervention; effective therapy; epigenome editing; excitatory neuron; gene repression; genetic variant; genome wide association study; genome-wide; genomic data; indexing; induced pluripotent stem cell; novel; risk prediction; risk variant; single-cell RNA sequencing; stem cell model; transcription factor

Alzheimer’s disease (AD) is a devastating neurodegenerative disorder without an effective cure. Recent genome- wide association studies (GWAS) on AD have identified >20 reproducible risk loci, providing an opportunity for understanding novel aspects of AD biology and developing effective interventions. However, each GWAS locus typically spans several genes and many equally associated genome-wide significant index/proxy single- nucleotide polymorphisms (SNPs) that are in strong linkage disequilibrium; it thus remains challenging to identify which genes and risk SNPs are causally involved in AD pathogenesis. Most GWAS risk variants are noncoding and likely regulate gene expression. Because accessible or open chromatin overlaps with cis-regulatory sequences, we hypothesize that many causal AD risk variants modulate chromatin accessibility to transcription factors, thereby altering molecular and cellular phenotypes relevant to AD. We have recently shown that open chromatin profiles in neurons derived from human induced pluripotent stem cells (hiPSCs) can help prioritize regulatory GWAS risk variants of schizophrenia. By directly comparing the quantitative measurements of open chromatin between the two alleles of a heterozygous SNP within the same sample, i.e., allele-specific open chromatin (ASoC) assay, we further showed that schizophrenia GWAS risk variants frequently exhibit ASoC in hiPSC-neurons. More importantly, we found that the neuronal ASoC variants are highly enriched for AD GWAS risk variants and can readily inform putatively regulatory risk variants at three leading AD risk loci (BIN1, CD2AP and CLU). Here, we will extend the ASoC approach to other hiPSC-derived AD-relevant cell types and also harness our expertise in CRISPR genome/epigenome editing and AD cell biology to address three specific questions: (1) Which AD GWAS risk variants are functional? For this, we will map open chromatin by ATAC-seq in glutamatergic and GABAergic neurons, astrocytes and microglia, and search for regulatory AD-risk variants that present ASoC; (2) Which genes are regulated by the putatively functional AD GWAS variants and are thus likely to be causal? For this, we will cost-effectively assay the regulatory effects of all putative AD-risk variants in hiPSC-derived neurons, astrocytes and microglia by combining multiplexed CRISPR/cas9 epigenome editing with single-cell RNA-seq; (3) What are the cellular phenotypic changes caused by the putatively causal AD variants/genes? For this, we will study the cis-regulatory effects of regulatory variants in BIN1, CD2AP and CLU (and other prioritized genes) on AD-relevant biochemical and cellular phenotypes in CRISPR-engineered hiPSC models. This project would impact the field by moving beyond GWAS to decipher causal mechanisms and develop effective treatments.

阿尔茨海默病（AD）是一种严重的神经退行性疾病，目前尚无有效的治疗方法。最近的基因组- 关于AD的广泛关联研究（GWAS）已经确定了>20个可重复的风险位点， 了解AD生物学的新方面并开发有效的干预措施。然而，每个GWAS位点 通常跨越几个基因和许多同样相关的全基因组显着指数/代理单- 核苷酸多态性（SNP）处于强烈的连锁不平衡;因此， 哪些基因和风险SNP与AD发病机制有因果关系。大多数GWAS风险变体是非编码的 并可能调节基因表达。因为可接近的或开放的染色质与顺式调节的 序列，我们假设许多致病性AD风险变体调节染色质对转录的可及性， 因子，从而改变与AD相关的分子和细胞表型。我们最近表明，开放 来源于人类诱导多能干细胞（hiPSC）的神经元中的染色质谱可以帮助优先考虑 精神分裂症的GWAS风险变异。通过直接比较开放的定量测量， 同一样品内杂合SNP的两个等位基因之间的染色质，即，等位基因特异性开放 我们进一步发现，精神分裂症GWAS风险变异体经常在精神分裂症患者中表现出ASoC。 hiPSC-神经元。更重要的是，我们发现神经元ASoC变体高度富集AD GWAS， 风险变异，并可以很容易地告知在三个主要的AD风险基因座（BIN 1，CD 2AP CLU）。在这里，我们将ASoC方法扩展到其他hiPSC衍生的AD相关细胞类型， 利用我们在CRISPR基因组/表观基因组编辑和AD细胞生物学方面的专业知识， 问题：（1）哪些AD GWAS风险变体是功能性的？为此，我们将通过ATAC-seq绘制开放染色质 在多巴胺能和GABA能神经元、星形胶质细胞和小胶质细胞中，并寻找调节AD风险的变体 （2）哪些基因受pupillary功能性AD GWAS变体的调节，因此 有可能是因果关系吗为此，我们将成本有效地分析所有推定的AD风险变体的调节作用 通过组合多重CRISPR/cas9表观基因组编辑， 用单细胞RNA-seq;（3）由脓疱性AD引起的细胞表型变化是什么 变异/基因？为此，我们将研究BIN 1、CD 2AP和CLU中调节变体的顺式调节作用 (and其他优先基因）对CRISPR工程化的hiPSC中AD相关生物化学和细胞表型的影响 模型该项目将通过超越GWAS来破译因果机制， 开发有效的治疗方法。