Understanding the Role of the Integrated Stress Response in tRNA Synthetase-associated Charcot-Marie-Tooth Disease

了解综合应激反应在 tRNA 合成酶相关夏科-马里-图思病中的作用

基本信息

批准号：
10740335
负责人：
Timothy Hines
金额：
$ 12.22万
依托单位：
JACKSON LABORATORY
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：
2023-08-01 至 2025-07-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10740335
关键词：
Affect Alleles Amino Acids Amino Acyl-tRNA Synthetases Autopsy Axon Binding Biological Models Career Transition Award Cell Fraction Cell Line Cells Charcot-Marie-Tooth Disease Charge Clustered Regularly Interspaced Short Palindromic Repeats Data Defect Demyelinations Disease Distal Drug usage EIF-2alpha Engineering Ensure Enzymes Event FGF21 gene Family Functional disorder GDF15 gene Gene Expression Gene Family Genes Genetic Engineering Goals Hereditary Disease Histology Human Human Engineering Immunofluorescence Immunologic In Vitro Inherited Knock-out Ligase Link Mammalian Genetics Measures Mediating Mentors Methods Modeling Morphologic artifacts Motor Motor Neurons Mus Mutation Nerve Degeneration Neuromuscular Diseases Neuromuscular Junction Neuronal Dysfunction Neuropathy Paper Pathogenesis Pathogenicity Pathology Pathway interactions Patients Peptide Initiation Factors Peripheral Nerves Peripheral Nervous System Diseases Persons Phase Phenotype Phosphorylation Phosphotransferases Physiology Postdoctoral Fellow Production Protein Biosynthesis Proteins Proteomics Publishing Quantitative Reverse Transcriptase PCR RNA Interference Rare Diseases Reagent Resource Development Review Literature Ribosomes Role Sampling Science Series Serum Skeletal Muscle Source Spinal Cord System Testing The Jackson Laboratory Therapeutic Therapeutic Intervention Tissue Sample Tooth Diseases Training Transfer RNA Transgenic Mice Translations Tyrosine-tRNA Ligase Up-Regulation Variant Western Blotting Work YARS gene afferent nerve biological adaptation to stress career career development causal variant data integration differential expression directed differentiation disease phenotype disease-causing mutation dominant genetic mutation experimental study fibroblast growth factor 21 gene replacement human disease human stem cells in vivo induced pluripotent stem cell mouse model mutant neuromuscular neuronal cell body novel therapeutics overexpression prevent protein expression response sensor stem cell model targeted treatment therapeutic evaluation therapeutic target tool transcription factor transcriptome sequencing transcriptomics

项目摘要

PROJECT SUMMARY This proposal will address the mechanisms underlying neuromuscular degeneration in Charcot-Marie-Tooth disease (CMT). CMT is a genetically and phenotypically heterogeneous neuromuscular disorder with causative mutations found in over 100 genes. While considered a rare disease, CMT is the most common inherited disorder of the peripheral nervous system, affecting ~1 in 3,500 people worldwide. Dominant mutations in 6 different tRNA synthetases (aaRSs) cause forms of CMT (aaRS-CMT), making them the largest family of CMT-associated genes. Each of these genes is involved in protein synthesis suggesting a common mechanism that leads to defects in protein production and ultimately CMT pathologies. Our recently published work uncovered a potential mechanism underlying aaRS-CMT. We found that mutant aaRSs inappropriately sequester tRNAs from the ribosome, which stalls ribosome function and activates an integrated stress response (ISR) via a sensor protein, GCN2. ISR activation causes two major cellular events: 1) shutdown of a major form of protein synthesis and, 2) upregulation of the transcription factor, ATF4, and its target genes. The relative contributions of each of these events is currently unknown. One goal of this project is to determine the role of ATF4 and target genes in the pathophysiology observed in aaRS-CMT. Preliminary results show that ATF4 overexpression is toxic to motor neurons and produces a CMT- like phenotype in mice, evidence that ATF4 could be a viable therapeutic target for aaRS-CMT. In Aim 1 we will manipulate ATF4 expression levels in validated mouse models of aaRS-CMT to determine whether the disease pathology is driven by decreased protein translation or by increased expression of the ATF4 gene. To advance toward therapeutic applications we need to establish that human motor neurons also activate the ISR in response to aaRS-CMT mutations. Therefore, in Aim 2 we will establish and validate human induced pluripotent stem cell (hiPSC)-derived motor neuron cultures which have been genetically engineered to model aaRS-CMT. We will also test therapeutic strategies in these human cell-based models. Interestingly, ATF4 expression is common in many different types of neurodegeneration. Therefore, in Aim 3, we will integrate data from ATF4 mice in Aim 1, and hiPSC-derived motor neurons in Aim 2 to identify common genes and cellular pathways involved in ATF4-mediated neurodegeneration. These hiPSC-based models will be a powerful tool to help identify and develop new targets or pathways for potential therapeutic interventions. This MOSAIC (Maximizing Opportunities for Scientific and Academic Independent Careers) Postdoctoral Career Transition Award to Promote Diversity will be supported by excellent career development resources and a mentoring team of globally recognized experts in CMT (R.W. Burgess) and human stem cells (M.F. Pera) at The Jackson Laboratory for Mammalian Genetics.

项目摘要该提案将解决charcot-marie-tooth中神经肌肉变性的基础机制疾病（CMT）。 CMT是一种遗传和表型异质性神经肌肉疾病，具有病因在100多个基因中发现的突变。虽然被认为是一种罕见疾病，但CMT是最常见的遗传疾病在周围神经系统中，全世界3500人中有约1人。 6种不同的tRNA中的主要突变合成酶（AARSS）引起CMT（AARS-CMT）的形式，使其成为CMT相关的最大家族基因。这些基因中的每一个都参与蛋白质合成，这表明一种常见机制，导致蛋白质产生和最终CMT病理的缺陷。我们最近发表的工作发现了AARS-CMT的潜在机制。我们发现该突变体 AARS不恰当地从核糖体中隔离TRNA，该核糖体阻滞了核糖体功能并激活通过传感器蛋白GCN2进行综合应力响应（ISR）。 ISR激活导致两个主要的细胞事件： 1）关闭主要形式的蛋白质合成和2）转录因子ATF4及其上调靶基因。这些事件中每个事件的相对贡献目前尚不清楚。该项目的一个目标是确定ATF4和靶基因在观察到的病理生理学中的作用 AARS-CMT。初步结果表明，ATF4过表达对运动神经元有毒，并产生CMT- 像小鼠的表型一样，证据表明ATF4可能是AARS-CMT的可行治疗靶标。在目标1中，我们将在经过验证的AARS-CMT小鼠模型中操纵ATF4表达水平，以确定该疾病是否是否病理是由蛋白质翻译降低或ATF4基因表达增加的驱动。为了迈向治疗应用，我们需要确定人类运动神经元也激活 ISR响应AARS-CMT突变。因此，在AIM 2中，我们将建立并验证人类诱导的多能干细胞（HIPSC）衍生的运动神经元培养物已经过基因设计以建模 AARS-CMT。我们还将在这些基于人类细胞的模型中测试治疗策略。有趣的是，ATF4 表达在许多不同类型的神经变性中很常见。因此，在AIM 3中，我们将集成数据来自AIM 1中的ATF4小鼠，AIM 2中的HIPSC衍生运动神经元以鉴定常见基因和细胞与ATF4介导的神经变性有关的途径。这些基于HIPSC的模型将是一个强大的工具帮助识别并开发潜在治疗干预措施的新目标或途径。这种马赛克（最大化科学和学术独立职业的机会）职业生涯促进多样性的过渡奖将得到出色的职业发展资源的支持和 CMT（R.W. Burgess）和人类干细胞（M.F. Pera）的全球认可专家的指导团队杰克逊哺乳动物遗传学实验室。