THE HUMAN BRAINOME III: EQTL REGULATION BY NATURAL ANTISENSE RNA IN ALZHEIMER S DISEASE

人类大脑 III：天然反义 RNA 对阿尔茨海默病的 EQTL 调节

• 批准号: 10256018
• 负责人: Amanda J Myers
• 金额: $ 77.13万
• 依托单位: UNIVERSITY OF MIAMI SCHOOL OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-09-15 至 2025-05-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10256018
• 关键词: Affect; Alleles; Alzheimer' s Disease; Alzheimer' s Disease Pathway; Alzheimer' s disease pathology; Amyloid beta-Protein; Antisense RNA; Archives; Binding; Biology; Brain; Cell Culture Techniques; Code; DNA; Data; Data Collection; Databases; Disease; Genomics; Genotype; Goals; Human; Institutes; International; Investigation; Laboratories; Late Effects; Late Onset Alzheimer Disease; Length; Maps; Measures; Oligonucleotides; Outcome; Output; Pathogenicity; Pathologic; Pathway interactions; Peptides; Procedures; Process; Proteins; Proteomics; Quantitative Trait Loci; RNA; Regulation; Reproducibility; Resources; Risk; Sampling; Series; Site; Specificity; Techniques; Technology; Testing; Time; Tissue Banks; Transcript; Translations; Untranslated RNA; Validation; Variant; Work; base; brain tissue; case control; differential expression; experimental study; genome wide association study; genomic variation; innovation; interest; knock-down; neurogenomics; novel; overexpression; risk variant; scaffold; single molecule; single molecule real time sequencing; tau Proteins; transcriptome sequencing; transcriptomics; web site

PROJECT SUMMARY Previously, we have taken an innovative approach (The Human Brainome; [1-6]) to mapping risk loci for late onset Alzheimer's disease (LOAD). Rather than looking at a single layer of information as in most genome- wide association (GWAS) studies, we have mapped genomic variation in the context of downstream transcriptomic and proteomic expression. This allows for mapping both the crucial variation involved in LOAD, as well as the downstream effects and their directions. Additionally, it allows for building networks of multiple players crucial for disease processes. One shortcoming of the current work is that we have mapped DNA-expression relationships that are subtly changed in Alzheimer's, but we have yet to fully understand why those DNA-expression relationships are altered. We know that expression is altered by specific alleles, but there must be added regulation given our mapped outputs. One target that can alter pathways are natural antisense transcripts (NATs), which can bind to oligonucleotide products and alter their expression and degradation. In our application, we propose to use long read sequencing technology (SMRT; Single Molecule, Real-Time) and fully profile RNA from our human brain bank samples. We will examine where these outputs are located and perform preliminary work to determine if any of these new hits can act on our existing results. We propose to follow these targets through 3 Aims. Aim 1 will involve following hits from public databases. Aim 2 will involve collecting additional RNA profiling data. Finally, Aim 3 will seek to validate and order all novel findings from Aims 1 and 2. It is important to use technologies appropriate to our hypothesis for the new data collection. The majority of non-coding RNA belongs to the class of transcripts called long non-coding RNA (lncRNA), which can span from 1000-10,000 bp [7]. Typical short-read RNA sequencing (SRS) technologies are based on the capture of short sequences of ~150 bp, and therefore, SRS has difficulty in capture and alignment of longer products. We are working with Robert Sebra at the Icahn Institute for Genomics and Multicale Biology, who is an expert in SMRT sequencing [8]. This technology offers longer read lengths and will be unique-in-field, since most human RNA profiling involves SRS. By the completion of these Aims, we will have 1. A map of novel long read sequencing in human brain tissues, which will be a significant add-to-field, given most technologies used to date are focused on short read sequencing, and there is limited profiling in pathologically defined human brain tissues, 2. An understanding of how these novel hits are affecting both the direct sense transcript of interest as well as the known LOAD pathologies, 3. Multi-level mapping of rigor and reproducibility of targets through the use of multiple capture techniques and 4. Validation of the effect of hits on the known LOAD pathogenic targets by measuring both expression and protein levels in cell culture.

项目总结 以前，我们采取了一种创新的方法(人脑基因组；[1-6])来绘制风险基因座图 迟发性阿尔茨海默病(LOAD)。而不是像大多数基因组那样只看一层信息- 广泛关联(GWAS)研究，我们已经绘制了下游背景下的基因组变异图 转录和蛋白质组表达。这允许映射在负载中涉及的关键变化， 以及下游的影响和方向。此外，它还允许构建多个 在疾病过程中起关键作用的球员。 当前工作的一个缺点是我们已经绘制了微妙的DNA-表达关系图 在阿尔茨海默氏症中发生了变化，但我们还没有完全理解为什么这些DNA-表达关系 被更改了。我们知道表达是由特定的等位基因改变的，但考虑到我们的 映射的输出。可以改变途径的一个靶点是天然的反义转录本(NAT)，它可以结合 寡核苷酸产物并改变其表达和降解。在我们的应用程序中，我们建议使用 长读测序技术(SMRT；单分子，实时)和完整的人类RNA图谱 脑库样本。我们将检查这些输出的位置，并执行初步工作，以 确定这些新的点击量中是否有任何一个可以在我们现有的结果上发挥作用。 我们建议遵循这些目标实现三个目标。目标1将涉及公共数据库中的以下点击量。 目标2将涉及收集额外的RNA图谱数据。最后，《目标3》将对所有小说进行验证和排序 来自目标1和目标2的发现。对于新数据，使用适合我们假设的技术是很重要的 收集。大多数非编码RNA属于被称为长非编码RNA的转录本类别 (LncRNA)，其跨度可从1000-10000个碱基对[7]。典型的短读RNA测序(SRS)技术 是基于捕获~150个碱基的短序列，因此，SRS很难捕获和 较长产品的对准。我们正在与伊坎基因组研究所的罗伯特·塞布拉合作 他是SMRT测序方面的专家[8]。这项技术提供了更长的读取长度和 在现场是独一无二的，因为大多数人类RNA图谱涉及SRS。 通过这些目标的完成，我们将有1.人脑中的小说长阅读测序图 纸巾，这将是一个重要的补充领域，因为迄今为止使用的大多数技术都集中在短读上 测序，在病理定义的人脑组织中有有限的侧写，2.对 这些小说是如何影响直接意义上的感兴趣的文本以及已知的负载的 病理学，3.通过使用多次捕获对目标的严密性和重复性进行多层次映射 技术和4.通过测量两种方法验证HITS对已知负载致病目标的影响 在细胞培养中的表达和蛋白质水平。