The tRNA pool in C9-ALS/FTD

C9-ALS/FTD 中的 tRNA 池

• 批准号: 10662716
• 负责人: Ya-Ming Hou
• 金额: $ 24.92万
• 依托单位: THOMAS JEFFERSON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-04-01 至 2025-03-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10662716
• 关键词: ALS patients; Address; Adult; Affect; Aging; Amino Acids; Amyotrophic Lateral Sclerosis; Animal Model; Anticodon; Award; Base Sequence; Biological Models; Biology; Brain; C9ORF72; Cell model; Cells; Charge; Chemicals; Chimeric Proteins; Codon Nucleotides; Cognitive; Data; Development; Dipeptides; Disease; Disease Outcome; Foundations; Frontotemporal Dementia; Genes; Genetic; Genetic Transcription; Initiator Codon; Introns; Length; Link; Mediating; Medical; Messenger RNA; Modeling; Modification; Neurodegenerative Disorders; Neuronal Differentiation; Nucleosides; Nucleotides; Onset of illness; Pathogenesis; Pathology; Patients; Positioning Attribute; Protein Biosynthesis; Proteins; RNA; Reading Frames; Regulation; Reporter; Research; Ribosomal Frameshifting; Ribosomal Proteins; Ribosomes; Role; Running; Sampling; Secure; Temporal Lobe; Testing; Therapeutic; Therapeutic Intervention; Time; Tissues; Transfer RNA; Transfer RNA Aminoacylation; Translating; Translation Initiation; Translations; United States National Institutes of Health; Work; bench to bedside; curative treatments; design; frontal lobe; frontier; frontotemporal lobar dementia amyotrophic lateral sclerosis; genome-wide analysis; improved; induced pluripotent stem cell; insight; interest; loss of function; methyl group; motor neuron degeneration; novel; novel therapeutics; posttranscriptional; programs

PROJECT SUMMARY The C9orf72-mediated ALS (amyotrophic lateral sclerosis) and FTD (frontotemporal dementia) (C9- ALS/FTD) are two fatal neurodegenerative diseases with no curative treatment. We aim to address a new mechanism at the core of C9-ALS/FTD to improve the therapeutic potential. While ALS is an adult-onset disease with progressive degeneration of motor neurons, and while FTD is characterized with progressive decline of the frontal and temporal lobes of the brain, the two share a common genetic cause – repeat expansion of the nucleotide sequence G4C2 in the first intron of the C9orf72 gene. Of the proposed causes of C9-ALS/FTD, the most significant is the synthesis of dipeptide repeat (DPR) proteins from ribosome translation of the expanded repeats. These DPRs are consistently observed in patient tissues and in various model systems. While the initiation of DPR synthesis is non-canonical, the elongation of protein synthesis is by the classical ribosome machinery that translates each codon using a set of tRNA species with the matching anticodon, which is provided by the cellular tRNA pool. Notably, translation of repeated sequences is highly challenging, requiring repeated use of the tRNAs for the same codon, where each species must be efficiently charged and post-transcriptionally modified and matured. This challenge is further intensified in patient cells, where the repeat length can run up to thousands, raising the question of how the cellular tRNA pool responds to such an unusual demand. This is an unexplored, but critical, question in C9-ALS/FTD, because the deficiency of quality tRNAs can shift the ribosome translational reading-frame, leading to frameshifting and synthesis of chimeric DPRs that would alter the disease pathology. Here we will address this question, based on our extensive expertise in tRNA and in tRNA-associated ribosome frameshifting. In Aim 1, we will determine changes of the tRNA pool in C9 patient-derived cells relative to an isogenic control, using our improved tRNA-seq. We will also decipher changes of the ribosome-mediated global protein synthesis that underlie the disease. We will determine changes both in the iPSC (induced pluripotent stem cell) state and in the differentiated neuron (iPSN) state, due to their fundamental differences in regulation of protein synthesis, and due to the potential to produce new insight into the disease during differentiation. In Aim 2, we will test the hypothesis that the course of DPR synthesis can be induced to undergo a desired frameshifting to produce less toxic proteins that can reduce the disease pathology. We will test how changes of the abundance and charging levels of the tRNAs required for DPR synthesis can induce the desired frameshifting. This work will serve as the foundation for understanding the therapeutic implications of tRNA and ribosome protein synthesis in the development of C9-ALS/FTD, providing a template that is generalizable to other aging and neurodegenerative diseases associated with nucleotide repeat expansions.

项目摘要 C9ORF72介导的ALS（肌萎缩性侧硬化）和FTD（额颞痴呆）（C9-- ALS/FTD）是两种致命的神经退行性疾病，没有治疗治疗。我们旨在解决新的 C9-ALS/FTD核心的机制，以提高治疗潜力。而ALS是一种成人疾病 运动神经元的进行性变性，而FTD的特征是 大脑的额叶和临时爱，两者有共同的遗传原因 - 重复扩展 C9ORF72基因的第一个内含子中的核苷酸序列G4C2。在提议的C9-ALS/FTD的原因中 最重要的是从扩展的核糖体翻译中合成二肽重复（DPR）蛋白 重复。这些DPR在患者组织和各种模型系统中始终观察到。而 DPR合成的启动是非典型的，蛋白质合成的伸长是经典核糖体 使用与匹配的反密码子一起使用一组tRNA物种翻译每个密码子的机械 通过细胞tRNA池。值得注意的是，重复序列的翻译非常具有挑战性，需要重复 将TRNA用于同一密码子，其中每个物种都必须有效地充电并在转录后充电 修改和成熟。该挑战进一步启发了患者细胞，重复长度可以延伸到 成千上万的问题提出了一个问题，即细胞tRNA池如何响应这种异常需求。这是一个 C9-ALS/FTD中出乎意料但至关重要的问题，因为质量TRNA的缺乏可以改变核糖体 翻译阅读框架，导致将改变疾病的嵌合DPR的框架和合成 病理。在这里，我们将根据我们在tRNA和tRNA相关方面的广泛专业知识来解决这个问题 核糖体框架。在AIM 1中，我们将确定C9患者衍生细胞相对的tRNA池的变化 使用我们改进的tRNA zeq进行等源控制。我们还将破译核糖体介导的变化 疾病构成的全球蛋白质合成。我们将确定IPSC中的更改（引起 多能干细胞）状态和分化的神经元（IPSN）状态，由于它们的基本差异 调节蛋白质合成，并且由于有潜力对疾病产生新的见解 分化。在AIM 2中，我们将检验以下假设，即DPR合成的过程可以诱导进行 所需的帧速率可产生较小的毒性蛋白，从而减少疾病病理。我们将测试如何 DPR合成所需的TRNA的抽象和充电水平的变化可以诱导所需的 框架。这项工作将成为理解tRNA的治疗意义的基础 C9-ALS/FTD开发中的核糖体蛋白质合成，提供了可推广到的模板 与核苷酸重复扩张相关的其他衰老和神经退行性疾病。