Role of dorsal root ganglion FTO, a RNA demethylase, in neuropathic pain

背根神经节 FTO（一种 RNA 去甲基酶）在神经性疼痛中的作用

• 批准号: 10470094
• 负责人: Yuan-Xiang Tao
• 金额: $ 50.99万
• 依托单位: RUTGERS BIOMEDICAL AND HEALTH SCIENCES
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-07-15 至 2024-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10470094
• 关键词: Absence of pain sensation; Affect; Animals; Attenuated; Binding; Cell Nucleus; Chronic; Code; Data; Development; Disease; Down-Regulation; Epigenetic Process; Fatty acid glycerol esters; Gene Expression; Gene Expression Regulation; Genes; Genetic; Genetic Transcription; Histones; Homologous Gene; Human; Hypersensitivity; Injury; Lead; Ligation; Link; Maintenance; Malignant Neoplasms; Messenger RNA; Modification; Morphine; Neurons; Neuropathy; Obesity; Opioid Analgesics; Pain; Peripheral nerve injury; Pharmacology; Pilot Projects; Prevention strategy; Proteins; Public Health; RNA; RNA Degradation; RNA metabolism; Rattus; Regulator Genes; Role; Spinal Anesthesia; Spinal Ganglia; Spinal nerve structure; Symptoms; Testing; Time; Transcription Coactivator; Transcriptional Activation; Up-Regulation; WT1 gene; central sensitization; chronic pain; dorsal horn; histone methyltransferase; human disease; injured; knock-down; mechanical allodynia; morphine tolerance; mu opioid receptors; nerve injury; neurotransmitter release; novel; novel therapeutics; opiate tolerance; overexpression; pain reduction; pain symptom; painful neuropathy; promoter; transcription factor; treatment strategy

PROJECT SUMMARY: RNA modifications were recently rediscovered as essential regulators of gene expression. N6-methyladenosine (m6A) is identified as the most prevalent internal modification of eukaryotic RNA. The m6A modification controls RNA metabolism including RNA degradation and has been linked to human diseases such as obesity and cancer. However, its role in chronic pain including neuropathic pain is unknown. Our preliminary data suggest that a nerve injury-induced increase in dorsal root ganglion (DRG) FTO (fat mass and obesity-associated protein), a well-characterized RNA demethylase, may participate in neuropathic pain by reducing the level of m6A on Ehmt2 mRNA (encoding G9a, a histone methyltransferase), stabilizing nerve injury-induced G9a upregulation, and, consequently, silencing the expression of mu opioid receptor (MOR) in the injured DRG. Given that the transcriptional and translational changes in DRG gene expression following peripheral nerve injury participate in the development and maintenance of neuropathic pain and that G9a as a repressor of gene expression is a key endogenous contributor to neuropathic pain genesis, we propose that the increased FTO in the injured DRG contributes to neuropathic pain. In Aim 1, we will first determine whether pharmacological inhibition or genetic knockdown of DRG FTO attenuates nerve injury-induced pain hypersensitivity during the development and maintenance periods, and whether mimicking nerve injury-induced increase in DRG FTO leads to major symptoms of neuropathic pain in naive rats. In Aim 2, we will examine whether and how peripheral nerve injury upregulates the expression of FTO in the DRG. Time-dependent changes in the expression of Fto and its transcriptional activator Runx1 mRNAs and their proteins and in the level of m6A on specific RNAs in the DRG after peripheral nerve injury will be examined. We will also define whether nerve injury-induced up-regulation of DRG FTO is attributed to an increase in Runx1 in the injured DRG. In Aim 3, we will examine how DRG FTO participates in neuropathic pain. We will first observe whether FTO binds to Ehmt2 mRNA and whether this binding activity is increased in the injured DRG neurons after the fifth lumbar spinal nerve ligation (SNL). We will then define whether FTO contributes to the SNL-induced upregulation of Ehmt1 mRNA/G9a and the G9a-controlled downregulation of MOR by stabilizing the increased G9a expression in the injured DRG. Finally, we will determine whether blocking the SNL-induced increase in DRG FTO reduces the MOR-controlled primary afferent neurotransmitter release, restores the decrease of opioid analgesia, and attenuates opioid tolerance development. These studies will not only advance our understanding of posttranscriptional mechanisms of neuropathic pain, but will also open a door to develop a new strategy for the prevention and treatment of this disorder.

项目摘要： 最近将RNA修饰重新发现为基因表达的基本调节剂。 N6-甲基腺苷 （M6A）被确定为真核RNA的最普遍的内部修饰。 M6A修改控件 RNA代谢包括RNA降解，并与肥胖症和人类疾病有关 癌症。但是，其在包括神经性疼痛在内的慢性疼痛中的作用尚不清楚。我们的初步数据暗示 神经损伤引起的背根神经节（DRG）FTO（脂肪质量和肥胖相关）增加 蛋白质）是一种特征良好的RNA脱甲基酶，可以通过降低水平来参与神经性疼痛 M6A在EHMT2 mRNA上（编码G9A，组蛋白甲基转移酶），稳定神经损伤诱导的G9a 上调，因此，在受伤的DRG中沉默了MU阿片受体（MOR）的表达。 鉴于外周神经后DRG基因表达的转录和翻译变化 受伤参与神经性疼痛的发展和维持，而G9a作为基因的阻遏物 表达是神经性疼痛创世纪的关键内源性贡献者，我们提出增加的FTO在 受伤的DRG导致神经性疼痛。在AIM 1中，我们将首先确定药理学是否是否 DRG FTO的抑制或遗传敲低可减轻神经损伤引起的疼痛性超敏反应 开发和维护期以及是否模仿神经损伤引起的DRG FTO增加 导致天真大鼠神经性疼痛的主要症状。在AIM 2中，我们将检查是否以及如何 周围神经损伤在DRG中上调FTO的表达。时间相关的变化 FTO及其转录激活剂Runx1 mRNA及其蛋白的表达以及M6A的水平 周围神经损伤后DRG中的特定RNA将被检查。我们还将定义是否神经 受伤引起的DRG FTO上调归因于受伤的DRG中RUNX1的增加。在AIM 3中，我们 将研究DRG FTO如何参与神经性疼痛。我们将首先观察FTO是否与EHMT2结合 mRNA以及第五腰脊柱后受伤的DRG神经元中这种结合活性是否增加 神经结扎（SNL）。然后，我们将定义FTO是否有助于SNL引起的EHMT1上调 mRNA/G9A和G9A控制的MOR的下调，通过稳定G9A的表达增加 受伤的DRG。最后，我们将确定阻止SNL诱导的DRG FTO的增加是否减少 由MOR控制的原发性传入神经递质释放，恢复阿片类镇痛的减少，并恢复 减弱阿片类药物耐受性的发展。这些研究不仅会提高我们对 神经性疼痛的转录后机制，但也将打开一扇门，以制定新的策略 预防和治疗这种疾病。