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.

肌萎缩侧索硬化症(ALS)是一种毁灭性和致命性的神经退行性疾病 以进行性肌肉无力和最终瘫痪为特征。肌萎缩侧索硬化症的已知危险因素有 年龄和男性不断增加；最近的数据还涉及服兵役，从而增加了 退伍军人管理局的医疗系统。大约90%的ALS病例被归类为散发性ALS(SALS)，没有 这种疾病的家族病史。其余病例以显性方式遗传，突变约30 目前已知导致肌萎缩侧索硬化症的基因(最常见的是C9orf72、SOD1、TARDBP、FUS)。 谷氨酸能传递的病理生理障碍引起的过度兴奋性 在所有类型的ALS病例中都有报道，这可能是一种常见的疾病途径，容易 运动神经元退化。已有研究表明，ALS的谷氨酸能(Glu)缺乏可能导致 由于GluA2 AMPA亚单位的RNA编辑效率低下，AMPA Glu受体的功能发生了变化。 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， 显示患者运动皮质中ADAR2下调和GluA2编辑效率低下 C9ORF72相关的ALS(C9 ALS)以及SALS患者脊髓运动神经元的表达。我们还报道了 ADAR2的下调及其在动物中几个靶点的编辑减少 遭受脊髓损伤(SCI)。此外，在脊髓损伤中，这些事件是由炎症反应触发的。 并导致基因表达的改变，这可能有助于脊髓损伤后运动神经元的过度兴奋。 根据这些数据以及脊髓损伤后ALS患者存在神经炎症的事实，我们推测 这：(1)ADAR2在C9 ALS和SALS中都下调，导致其靶标的RNA编辑发生变化。 特别是，除了GluA2，ADAR2还编辑其他对神经元很重要的分子 兴奋性[如GluA3-4、海人酸受体亚基GluK1-2、5-HT2CR以及 C9 ALS和SALS的钾(Kv1.1)和钙(Cav1.3)通道也发生改变；(2)下调 ADAR2在肌萎缩侧索硬化症中的作用是由几种与疾病相关的不同病理特征引起的。鉴于 SALS中ADAR2的减少是由神经炎症、功能性ADAR2在C9中的可用性所触发的 肌萎缩侧索硬化症还受到其细胞质滞留的影响，这是由于核质中断引起的。 运输。 我们将通过表征ADAR2中与肌萎缩侧索硬化相关的改变(1)来检验这些假设 表达和定位以及(2)ADAR2靶向转录本的编辑。这些研究将 对从C9和散发性C9患者尸检组织中获得的神经元进行激光显微解剖 ALS(Aim1)和人诱导多能干细胞(HiPSC)分化成的运动神经元 来源于ALS患者(AIM2)。此外，先前发表的与肌萎缩侧索硬化症相关转录的研究 改变使用了整个(细胞异质)组织标本。然而，由于 中枢神经系统细胞类型的异质性，无法从以下方面推断ALS转录组的准确评估 从这些样本中获得的数据，结合了来自所有细胞类型的信号。因此，我们将研究细胞- C9 ALS和SALS的类型特异性转录本(Aim3)。这些研究是通过新的方法实现的 最近在我们实验室开发的，应该会增加阐明疾病相关变异的可能性， 包括与ADAR2和编辑有关的内容。 总体而言，这是一项建立在我们的协作的跨学科性质基础上的创新建议 团队调查具有ALS特征的RNA编辑变化，并定义仅在 C9或散发性ALS。我们的目标是推进当前的知识，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