Metabolic perturbations as a modifier of repeat-associated non-AUG translation in C9orf72-linked ALS/FTD

代谢扰动作为 C9orf72 相关 ALS/FTD 中重复相关非 AUG 翻译的修饰剂

基本信息

批准号：
10271263
负责人：
Andrew Nelson
金额：
$ 4.6万
依托单位：
THOMAS JEFFERSON UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-09-17 至 2022-09-16
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10271263
关键词：
Address Amino Acids Amyotrophic Lateral Sclerosis Astrocytes Autopsy Bacterial Artificial Chromosomes Behavioral Biochemical Biological Models Blood - brain barrier anatomy Brain C9ORF72 Cell model Cells Cellular Metabolic Process Cellular biology Cessation of life Chronic Cognitive Detection Dipeptides Disease Disease model Energy Metabolism Energy-Generating Resources Engineering Enzyme-Linked Immunosorbent Assay Eukaryotic Initiation Factor-2 Event Exhibits Genes Genetic Glucose Goals Human Immunofluorescence Immunologic Impairment In Vitro Induced pluripotent stem cell derived neurons Initiator Codon Insulin Intervention Introns Knowledge Lead Link Mediating Metabolic Metabolic Pathway Mitochondria Modeling Molecular Molecular Biology Motor Motor Neurons Mus Mutation Nerve Degeneration Neurodegenerative Disorders Neurons PERK kinase Pathogenesis Pathogenicity Pathology Pathway interactions Patients Pharmacology Phenotype Phosphorylation Production Proteins Reporter Reporting Research Respiratory Chain Role SLC2A1 gene Spinal Cord Symptoms Techniques Testing Tissues Transgenic Mice Transgenic Organisms Translating Translations Viral Vector base biological adaptation to stress cognitive function deprivation detection method disease mechanisms study disease phenotype endoplasmic reticulum stress energy balance frontotemporal lobar dementia-amyotrophic lateral sclerosis glucose metabolism glucose transport immunocytochemistry in vivo in vivo Model induced pluripotent stem cell interest knock-down live cell imaging mortality risk mouse model neuron loss neurotoxic new therapeutic target novel therapeutics nutrient metabolism oxidative damage protein degradation respiratory solute stressor tool

项目摘要

PROJECT SUMMARY Metabolic perturbations as a modifier of repeat-associated non-AUG translation in C9orf72-linked ALS/FTD The -(GGGGCC)n- hexanucleotide repeat expansion (HRE) occurring in the C9orf72 (C9) gene is the most common cause of ALS and FTD and leads to the production of neurotoxic dipeptide repeat proteins (DPRs) by a noncanonical mechanism known as repeat-associated non-AUG (RAN) translation. Activation of the integrated stress response by a number of different cellular stressors has been shown to increase the occurrence of RAN translation without increasing the canonical AUG-driven translation. The core event in the ISR activation is phosphorylation of the α subunit of eukaryotic initiation factor 2 (eIF2α) by one of four kinases: PERK, PKR, GCN2, and HRI, each of which can be activated by different cellular stressors. I propose to address the role of metabolic perturbations and their potential modifier effect on C9-linked RAN translation. This could occur via activation of the ISR, specifically through the GCN2 and PERK pathways, which are activated by amino acid deprivation and ER stress, respectively. This is particularly relevant to understanding C9-ALS/FTD pathogenic mechanisms because a variety of metabolic perturbations, including hypermetabolism, glucose/energy deficiency, and alterations to the mitochondrial respiratory chain, are observed in ALS and could lead to ISR activation. For instance, glucose deficiency in neurons could force cells to metabolize amino acids for energy, potentially causing a deficit in amino acid (a.a.) supply and activating the ISR via GCN2. Likewise, perturbations to the respiratory chain could cause excessive ROS formation, leading to ER stress and activating the ISR via PERK. I propose to test the hypothesis that in C9- ALS/FTD, disease-driven energy imbalances promote RAN translation via ISR activation. I will employ both in vitro and in vivo models of the disease, including cortical and motor neurons differentiated from C9 patient-derived induced pluripotent stem cells (C9-iPSCs) as well as a C9orf72 bacterial artificial chromosome (BAC) transgenic mouse model. I will perturb the energy balance in these models by targeting critical metabolic pathways. Then I will evaluate the impact on ISR activation and RAN translation using biochemical, molecular and cell biology techniques and several behavioral analyses.

项目摘要代谢扰动作为C9ORF72链接ALS/FTD中重复相关非AUG翻译的修饰符 - （GGGGCC）N-六核苷酸重复扩展（HRE）发生在C9orf72（C9）基因中是最常见的原因 ALS和FTD的作品，并导致通过非规范机制产生神经毒性二肽重复蛋白（DPRS）称为重复相关的非aug（ran）翻译。通过许多已经显示出不同的细胞应激源可增加RAN翻译的发生而不增加规范 8月驱动的翻译。 ISR激活中的核心事件是真核开始的α亚基的磷酸化因子2（EIF2α）通过四个激酶之一：PERK，PKR，GCN2和HRI，每个激酶都可以通过不同的细胞激活。压力源。我建议解决代谢扰动的作用及其对C9连接RAN的潜在修饰效应翻译。这可能通过激活ISR，特别是通过GCN2和PERK途径的激活发生，分别被氨基酸剥夺和ER应激激活。这与了解C9-ALS/FTD特别相关致病机制，因为多种代谢扰动，包括多种代谢，葡萄糖/能量在ALS中观察到缺乏和线粒体呼吸链的改变，并可能导致ISR激活。为了例如，神经元中的葡萄糖缺乏症可能迫使细胞代谢氨基酸以获取能量，并可能导致赤字在氨基酸（A.A.）中，通过GCN2供应和激活ISR。同样，对呼吸链的扰动可能会导致过多的ROS形成，导致ER应力并通过PERK激活ISR。我建议检验以下假设。 ALS/FTD，疾病驱动的能量不平衡通过ISR激活促进翻译。我将在体外和该疾病的体内模型，包括与C9患者衍生的诱导的皮质和运动神经元多能干细胞（C9-IPSC）以及C9orf72细菌人造染色体（BAC）转基因小鼠模型。我通过靶向关键代谢途径，将在这些模型中扰动能量平衡。然后我将评估对 ISR激活并使用生化，分子和细胞生物学技术以及几种行为进行翻译分析。