The role of ADAR2-associated RNA editing in pathogenesis of ALS

ADAR2 相关 RNA 编辑在 ALS 发病机制中的作用

• 批准号: 10084222
• 负责人: STELLA DRACHEVA
• 金额: --
• 依托单位: JAMES J PETERS VA MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-10-01 至 2021-09-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10084222
• 关键词: ALS patients; AMPA Receptors; Acids; Age; Amyotrophic Lateral Sclerosis; Animals; Astrocytes; Autopsy; Biological Assay; Brain region; C9ORF72; CRISPR/Cas technology; Calcium; Cells; Coculture Techniques; DRADA2b protein; Data; Disease; Disease Pathway; Disease Progression; Down-Regulation; Enzymes; Event; Family history of; Fluorescence-Activated Cell Sorting; Gene Expression; Genes; Glutamates; Goals; Healthcare Systems; Heterogeneity; Human; Immunohistochemistry; Inflammatory; Inflammatory Response; Inherited; Kainic Acid Receptors; Knowledge; Laboratories; Lasers; Massive Parallel Sequencing; Measures; Mediating; Messenger RNA; Methods; Microglia; Molecular; Motor Cortex; Motor Neurons; Muscle Weakness; Mutation; Nature; Neurodegenerative Disorders; Neurons; Nuclear Antigens; Occipital lobe; Oligodendroglia; Paralysed; Pathogenesis; Pathologic; Patients; Population; Potassium; Process; Publishing; Quality of life; RNA Editing; Reporting; Research; Risk Factors; Role; Serotonin Receptor 5-HT2C; Signal Transduction; Specimen; Spinal; Spinal Cord; Spinal cord injury; System; Testing; Time; Tissues; Transcript; Transcription Alteration; Variant; Veterans; base; cell type; cytokine; differential expression; improved; induced pluripotent stem cell; innovation; male; military service; neuroinflammation; neuronal excitability; novel; nucleocytoplasmic transport; receptor; restriction enzyme; sex; sporadic amyotrophic lateral sclerosis; stem; superoxide dismutase 1; transcriptome; transcriptome sequencing; transmission process

Amyotrophic Lateral Sclerosis (ALS) is a devastating and fatal neurodegenerative disease that is characterized by progressive muscle weakness and eventual paralysis. The known risk factors for ALS are increasing age and male sex; recent data also implicate military service, thus putting an additional burden on the VA healthcare system. Approximately 90% of all ALS cases are classified as sporadic ALS (sALS) with no family history of the disease. The remaining cases are inherited in a dominant fashion, and mutations in ~30 genes (most commonly in C9orf72, SOD1, TARDBP, FUS) are currently known to cause ALS. Hyperexcitability that stems from pathophysiological disturbance of glutamatergic transmission has been reported in all types of ALS cases, and may be a common disease pathway that predisposes motoneurons to degeneration. It has been suggested that the glutamatergic (Glu) deficit in ALS may result from altered function of the AMPA Glu receptors due to inefficient RNA editing of the GluA2 AMPA subunit. The GluA2 editing is catalyzed by the editing enzyme ADAR2. Our data, as well as data published by others, showed the downregulation of ADAR2 and inefficient editing of the GluA2 in the motor cortex of patients with C9orf72-associated ALS (C9 ALS) as well as in spinal motoneurons of patients with sALS. We also reported the downregulation of ADAR2 and the consequent decrease of editing in several of its targets in animals subjected to spinal cord injury (SCI). Moreover, in SCI these events are triggered by inflammatory response and result in changes in gene expression that are likely to contribute to post-SCI motoneuron hyperexcitability. Based on these data and the fact that, as in SCI, neuroinflammation is present in ALS, we hypothesize that: (1) ADAR2 is downregulated in both C9 ALS and sALS, leading to alterations in RNA editing in its targets. In particular, in addition to GluA2, ADAR2 editing of other molecules which are important for neuronal excitability [such as, GluA3-4, kainate receptor subunits GluK1-2, serotonin 2C receptor (5-HT2CR), as well as potassium (Kv1.1) and calcium (Cav1.3) channels] is also altered in C9 ALS and sALS; (2) The downregulation of ADAR2 in ALS results from several different pathological features associated with the disease. Whereas the decrease of ADAR2 in sALS is triggered by neuroinflammation, the availability of the functional ADAR2 in C9 ALS is also influenced by its cytoplasmic retention, which is caused by a disruption of nucleocytoplasmic transport. We will test these hypotheses by characterizing ALS-associated alterations (1) in the ADAR2 expression and localization and (2) in editing of the transcripts that are targeted by ADAR2. These studies will be performed in laser microdissected neurons obtained from autopsy tissues of patients with C9 and sporadic ALS (Aim1) as well as in motoneurons differentiated from human induced pluripotent stem cells (hiPSC) derived from ALS patients (Aim2). In addition, previously published studies of ALS-associated transcriptional alterations employed whole (cellular heterogeneous) tissue specimens. However, because of the heterogeneity of cell types in the CNS, accurate assessment of the ALS transcriptome cannot be inferred from the data obtained from these specimens that combine signals from all cell types. Therefore, we will study cell- type-specific transcriptomes in C9 ALS and sALS (Aim3). These studies are enabled by the novel methods recently developed in our lab, and should enhance the likelihood of elucidating disease relevant variations, including those related to ADAR2 and editing. Overall, this is an innovative proposal that builds on the interdisciplinary nature of our collaborative team to investigate RNA editing changes that characterize ALS and to define unique changes occurring only in C9 or sporadic ALS. Our goal is to advance the current knowledge and to discover new treatments.

肌萎缩性外侧硬化症（ALS）是一种毁灭性和致命的神经退行性疾病 以进行性肌肉无力和最终瘫痪为特征。 ALS的已知风险因素是 年龄和男性的增长；最近的数据也暗示了兵役，从而增加了负担 VA医疗保健系统。大约90％的ALS案例被归类为零星ALS（SALS），没有 疾病的家族史。其余案例是以主导方式继承的，在〜30中的突变是 目前已知基因（最常见于C9orf72，SOD1，TARDBP，FUS）引起ALS。 源自谷氨酸能传播的病理生理障碍的过度兴奋性具有 在所有类型的ALS病例中都有报道，可能是一种常见的疾病途径 运动神经元变性。有人提出可能导致ALS中的谷氨酸能（GLU）缺陷 由于GLUA2 AMPA亚基的RNA编辑效率低下而导致AMPA GLU受体的功能的改变。 GLUA2编辑是由编辑酶ADAR2催化的。我们的数据以及其他人发布的数据 显示了ADAR2的下调以及GLUA2在运动皮层中的效率低下 C9ORF72相关的ALS（C9 ALS）以及SALS患者的脊髓运动神经元。我们还报告了 ADAR2的下调及其在动物中的几个目标中的编辑减少 遭受脊髓损伤（SCI）。此外，在SCI中，这些事件是由炎症反应触发的 并导致基因表达的变化，可能导致SCI后运动神经元过度兴奋性。 基于这些数据以及与SCI一样的事实，ALS中存在神经炎症，我们假设 那是：（1）ADAR2在C9 ALS和SALS中都被下调，导致其目标中RNA编辑的改变。 特别是，除了GLUA2之外，其他分子的ADAR2编辑对神经元很重要 兴奋性[例如，GluA3-4，海藻酸盐受体亚基GLUK1-2，5-羟色胺2C受体（5-HT2CR）以及 在C9 ALS和SALS中，钾（KV1.1）和钙（Cav1.3）通道也发生了变化。 （2）下调 ALS中的ADAR2的结果是与疾病相关的几种不同的病理特征。而 sals中ADAR2的降低是由神经炎症触发的，C9中功能性ADAR2的可用性 ALS还受其细胞质保留的影响，这是由于核质的破坏引起的 运输。 我们将通过表征ADAR2中与ALS相关的变化（1）来检验这些假设 在ADAR2靶向的转录本编辑中的表达和定位和（2）。这些研究会 在从C9患者尸检组织中获得的激光微解析神经元进行 ALS（AIM1）以及与人类诱导的多能干细胞（HIPSC）区分开的运动神经元（HIPSC） 源自ALS患者（AIM2）。此外，先前发表的有关ALS相关转录的研究 使用整体（细胞异质）组织样品的改变。但是，由于 中枢神经系统中细胞类型的异质性，无法从ALS转录组进行准确评估 从这些标本中获得的数据结合了所有细胞类型的信号。因此，我们将研究细胞 C9 ALS和SALS中的特定类型转录组（AIM3）。这些研究是通过新方法启用的 最近在我们的实验室中开发的，应加强阐明疾病相关变化的可能性， 包括与ADAR2和编辑有关的那些。 总体而言，这是一个创新的建议，基于我们协作的跨学科性质 团队调查RNA编辑更改的团队，这些更改的特征是ALS并定义仅发生的独特变化 C9或零星的ALS。我们的目标是促进当前的知识并发现新的治疗方法。

项目成果

Divergent single cell transcriptome and epigenome alterations in ALS and FTD patients with C9orf72 mutation.

• DOI: 10.1038/s41467-023-41033-y
• 发表时间: 2023-09-15
• 期刊: NATURE COMMUNICATIONS
• 影响因子: 16.6
• 作者: Li, Junhao;Jaiswal, Manoj K.;Chien, Jo-Fan;Kozlenkov, Alexey;Jung, Jinyoung;Zhou, Ping;Gardashli, Mahammad;Pregent, Luc J.;Engelberg-Cook, Erica;Dickson, Dennis W.;Belzil, Veronique V.;Mukamel, Eran A.;Dracheva, Stella
• 通讯作者: Dracheva, Stella