A cell model of YARS2-associated childhood-onset mitochondrial disease

YARS2 相关的儿童期发病线粒体疾病的细胞模型

基本信息

批准号：
10575369
负责人：
Ya-Ming Hou
金额：
$ 8.99万
依托单位：
THOMAS JEFFERSON UNIVERSITY
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：
2023-02-01 至 2025-01-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10575369
关键词：
Address Affect Alleles Amino Acids Amino Acyl-tRNA Synthetases Aminoacylation Award Biological Assay Cell Line Cell Signaling Process Cell model Central Nervous System Charge Childhood Clinical Clustered Regularly Interspaced Short Palindromic Repeats Codon Nucleotides Communities Complex Cytosol DNA Sequence Alteration Data Dedications Dermal Development Disease Disease Progression Enzymes Failure to Thrive Family Fibroblasts Foundations Functional disorder Gene Expression Genes Genetic Genotype Health Health Care Costs Heterozygote Homeostasis Human Human Cell Line Impairment Individual Intellectual functioning disability Joints Lactic Acidosis Link Manuscripts Mediating Methods Mitochondria Mitochondrial Diseases Mitochondrial Proteins Modeling Motor Neurons Muscle Cells Mutation Myopathy Neonatal Neuromuscular Diseases Neurons Nuclear Family Nutrient Organ Ovarian Pathogenicity Patients Peripheral Phenotype Positioning Attribute Protein Biosynthesis Public Health Publishing Pulmonary Hypertension Recurrence Research Role Secure Sensorineural Hearing Loss Severities Severity of illness Shapes Sideroblastic Anemia Signal Transduction Skeletal Muscle Speed Spinal Stem Cell Research Testing Tissue Model Tissues Transfer RNA Transfer RNA Aminoacylation Translations Tyrosine-Specific tRNA Tyrosine-tRNA Ligase United States National Institutes of Health Variant Vertebral column Work autosome bench to bedside cell type clinical diagnosis clinical phenotype clinically relevant detection of nutrient disease phenotype emerging adult enzyme activity established cell line frontier human disease human subject induced pluripotent stem cell infancy insight loss of function mutation correction myopathy-lactic acidosis-sideroblastic anemia syndrome neonate novel prognostication skeletal stem cell differentiation stem cell model transcriptome transcriptome sequencing

项目摘要

PROJECT SUMMARY We hypothesize that pathogenic variants in the family of the nuclear ARS2 genes, encoding mitochondrial aminoacyl-tRNA synthetases (mt-aaRSs), cause disease primarily by disrupting nutrient sensing and cell signaling. Each mt-aaRS is responsible for charging its cognate mitochondrial tRNA (mt-tRNA) with its specific amino acid, as required for the dedicated mitochondrial protein synthesis machinery. Loss-of-function variants of each ARS2 have been linked to human diseases, showing central nervous system involvement, myopathy, sensorineural hearing loss, ovarian dysgenesis, and pulmonary hypertension. However, the correlation is poor between the enzymatic activity of individual variants and manifestation of the disease phenotypes. This suggests that disease is mediated not necessarily via loss of tRNA charging, but by other functions of mt-aaRSs. Given that the cytosolic counterpart ARS1 genes are often implicated in non-canonical functions, beyond aminoacylation, we will test the hypothesis that ARS2s mediate nutrient-sensing and cell-signaling processes that are disrupted by pathogenic variants resulting in disease development. To test this hypothesis, as a pilot model, we will generate a human subject-derived induced pluripotent stem cell (iPSC) line with pathogenic variants in the YARS2 gene, encoding mt-TyrRS. Pathogenic variants of YARS2 result in the MLASA (myopathy, lactic acidosis and sideroblastic anemia) syndrome. Previous studies of pathogenic YARS2 variants have shown no appreciable effect on tRNA aminoacylation. Here we focus on a pair of novel compound heterozygous mutations that we identified in a human subject with neonatal fatal disease, the most severe clinical case observed to date. This provides a unique model where the largest changes in nutrient sensing and cell signaling are expected to occur. In Aim 1, we will create an iPSC line from an established fibroblast line of the human subject. We will use CRISPR/Cas to generate an isogenic control line that corrects the genetic mutations. In Aim 2, we will differentiate the patient and control iPSC lines to disease-relevant cell types. We will focus on myocytes, as skeletal myopathy is the most notable feature of clinical cases of YARS2 variants. We will also focus on peripheral neurons, which are affected in most clinical cases of other ARS2 variants. We will identify changes of gene expression in nutrient sensing or cell signaling of the subject line. Combined, this work will produce an isogenic pair of subject and control iPSC lines that will be shared with the research community. This pair of iPSC lines will serve as the foundation for understanding the implications of nutrient sensing and cell signaling in YARS2 variants, which will provide a template that is generalizable to other ARS2-associated disease.

项目摘要我们假设核ARS2基因家族中的致病变异，编码线粒体氨基酰基-TRNA合成酶（MT-AARSS），主要通过破坏营养感和细胞引起疾病信号。每个mt-a均负责为其同源线粒体tRNA（mt-tRNA）收取特定氨基酸，按专用线粒体蛋白合成机制所需的要求。功能丧失变体每个ARS2都与人类疾病有关，显示中枢神经系统的参与，肌病，感官听力损失，卵巢发育不全和肺动脉高压。但是，相关性很差在个体变体的酶活性和疾病表型的表现之间。这暗示着该疾病不一定是通过损失tRNA充电而介导的，而是通过MT-AARS的其他功能来介导的。给出胞质对应物ARS1基因通常与非规范功能有关，超出氨基酰化，我们将测试ARS2S介导营养感应和细胞信号过程的假设受到致病性变异的破坏，导致疾病发展。为了检验这一假设，作为飞行员模型，我们将生成具有致病性的人类受试者诱导的多能干细胞（IPSC）系 YARS2基因中的变体，编码MT-Tyrr。 YARS2的致病变异导致MLASA（肌病，乳酸酸中毒和副细胞贫血）综合征。病原Yars2变体的先前研究已显示对tRNA氨基酰化没有明显的影响。在这里，我们专注于一对新颖的复合杂合子我们在具有新生儿致命疾病的人类受试者中发现的突变，这是最严重的临床病例迄今为止观察到。这提供了一个独特的模型，其中营养感应和细胞信号的最大变化预计会发生。在AIM 1中，我们将从人类的既定成纤维细胞线创建IPSC线主题。我们将使用CRISPR/CAS生成纠正遗传突变的等源控制线。目标 2，我们将将患者和控制IPSC线与与疾病相关的细胞类型区分开。我们将专注于肌细胞，因为骨骼肌病是YARS2变体临床病例的最显着特征。我们还将专注于周围神经元，在其他ARS2变体的大多数临床病例中都受到影响。我们将确定受试者线的营养感应或细胞信号传导中的基因表达。合并，这项工作将产生与研究界共享的等源性主体和控制IPSC线路。这对IPSC 线条将成为理解营养感应和细胞信号传导含义的基础 YARS2变体将提供一个可推广到其他与ARS2相关疾病的模板。