DMA-Tudor interaction modules: a novel approach to Survival Motor Neuron protein (SMN) and Cajal body function

DMA-Tudor 相互作用模块：运动神经元生存蛋白 (SMN) 和 Cajal 身体功能的新方法

• 批准号: 10502150
• 负责人: Karla M Neugebauer
• 金额: $ 44.17万
• 依托单位: YALE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-07-15 至 2027-06-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10502150
• 关键词: Affect; Affinity; Amino Acids; Arginine; Axon; Binding; Binding Proteins; Binding Sites; Biochemistry; Biological Assay; Biotinylation; Cell Nucleus; Cell physiology; Cells; Cellular Stress; Chemicals; Complement; Cytoplasm; Data; Diffuse; Disease; Embryo; Embryonic Development; Etiology; Genetic; Genetic Transcription; Goals; Health; Image; Infant; Ligand Binding; Ligands; Location; Mass Spectrum Analysis; Mediating; Methylation; Microscopy; Missense Mutation; Modification; Molecular; Motor; Motor Neurons; Mus; Muscle; Mutation; Neurons; Nuclear; Optics; Patients; Phenotype; Physical condensation; Play; Positioning Attribute; Post-Translational Protein Processing; Protein Deficiency; Proteins; Publishing; RNA; RNA Processing; Role; SMN protein (spinal muscular atrophy); Site; Source; Specific qualifier value; Specificity; Spinal Muscular Atrophy; Stress; Structure; Testing; Time; Tissues; Toddler; Translations; Zebrafish; arginine methyltransferase; axonal pathfinding; dimethylarginine; fluorescence imaging; genome editing; globular protein; grasp; inhibitor; insight; mortality; mutant; nervous system disorder; neuromuscular system; novel; novel strategies; novel therapeutic intervention; protein complex; protein function; scaffold; tissue/cell culture

Survival Motor Neuron protein (SMN) deficiencies cause Spinal Muscular Atrophy (SMA), the most common genetic cause of infant and toddler mortality. Although SMN has been implicated in transcription, RNA processing and translation, the molecular basis of motoneuron loss is still unknown. We recently discovered a novel activity of SMN: biomolecular condensation caused by SMN’s globular tudor domain (SMNTud), which binds ligands modified by dimethylarginine (DMA). Although there are hundreds of DMA-modified proteins in cells, the correspondence between tudor domains and their DMA ligands remains unknown. This understudied post-translational modification is potentially dynamic and has emerging roles in multiple neurological diseases through the altered functions of cellular compartments known as biomolecular condensates (BMCs). Our central hypothesis is SMNTud binding to DMA ligands plays critical roles in cellular organization, which are especially vulnerable in the neuromuscular system. Our findings highlight a critical need to comprehensively determine the DMA ligands of SMNTud and the activities of the interaction modules they form. SMN is diffusely cytoplasmic and present in nuclear BMCs called Cajal bodies, which are essential for embryonic development. Cajal bodies are scaffolded by a known SMNTud ligand and altered in SMA. During stress, SMN forms BMCs in the cytoplasm. Our preliminary results directly implicate DMA binding and biomolecular condensation in SMA, because SMNTud activity was blocked by a single amino acid mutation, E134K, that blocks binding to DMA ligands and causes SMA. Chemical inhibitors of DMA drastically altered the composition and substructure of Cajal bodies, which we determined for the first time with our collaborator and co-I, Dr Joerg Bewersdorf. The identities of the full complement of DMA ligands that bind SMNTud and those that affect Cajal bodies have been unknown until now. Our preliminary data reveal ~70 novel and specific SMNTud ligands, indicating that new insights relevant to SMA are within our grasp. The overall objectives of this new application are to (i) identify the DMA ligands of SMNTud that mediate biomolecular condensation, (ii) understand the dynamicity of asymmetric (aDMA) and symmetric (sDMA) installed by arginine methyltransferases and removed by demethylases with respect to the structure and function of BMCs, and (iii) reveal novel SMN functions by leveraging DMA-SMNTud interaction modules. Our rationale is that objectives concerning DMA-SMNTud interaction modules will be most accessible to rigorous analysis through a combination of biochemistry and state-of-the-art imaging, using mouse and zebrafish cells and tissues, including motoneurons, with which we have expertise. If achieved, our aims will discover novel SMN binding partners and functions of SMNTud in biomolecular condensation. The regulatory potential and dynamicity of the DMA modification, a source of tissue specificity and disease etiology, will be determined. Identification of DMA-SMNTud interaction modules will suggest new therapeutic approaches for SMA.

运动神经元生存蛋白 (SMN) 缺乏会导致脊髓性肌萎缩症 (SMA)，这是最常见的疾病 婴儿和幼儿死亡的遗传原因。尽管 SMN 与转录有关，但 RNA 加工和翻译，运动神经元损失的分子基础仍然未知。我们最近发现了一个 SMN 的新颖活性：由 SMN 的球状 tudor 结构域（SMNTud）引起的生物分子缩合， 结合二甲基精氨酸 (DMA) 修饰的配体。尽管有数百种 DMA 修饰的蛋白质 在细胞中，tudor 结构域与其 DMA 配体之间的对应关系仍然未知。这 未被充分研究的翻译后修饰具有潜在的动态性，并且在多个领域发挥着新兴作用 通过改变被称为生物分子的细胞区室的功能而导致神经系统疾病 冷凝物（BMC）。我们的中心假设是 SMNTud 与 DMA 配体的结合在细胞中发挥着关键作用 组织，在神经肌肉系统中特别脆弱。 我们的研究结果强调了全面确定 SMNTud 的 DMA 配体及其活性的迫切需要 它们形成的交互模块。 SMN 广泛存在于细胞质中，存在于称为 Cajal 的核 BMC 中 体，对于胚胎发育至关重要。 Cajal 小体由已知的 SMNTud 配体支撑 并在 SMA 中发生改变。在应激期间，SMN 在细胞质中形成 BMC。我们的初步结果直接 暗示 SMA 中的 DMA 结合和生物分子缩合，因为 SMNTud 活性被 单氨基酸突变 E134K 可阻止与 DMA 配体的结合并导致 SMA。化学抑制剂 DMA 极大地改变了卡哈尔体的组成和子结构，这是我们第一次确定的 与我们的合作者兼同事 Joerg Bewersdorf 博士共度时光。完整 DMA 配体的身份 迄今为止，结合 SMNTud 的物质和影响 Cajal 小体的物质尚不清楚。我们的初步数据显示 约 70 种新颖且特定的 SMNTud 配体，表明与 SMA 相关的新见解就在我们的掌握之中。 这一新应用的总体目标是 (i) 识别介导 SMNTud 的 DMA 配体 生物分子缩合，(ii) 了解不对称 (aDMA) 和对称 (sDMA) 的动态性 由精氨酸甲基转移酶安装并由去甲基化酶去除的结构和 (iii) 通过利用 DMA-SMNTud 交互模块揭示新颖的 SMN 功能。我们的 基本原理是，有关 DMA-SMNTud 交互模块的目标将最容易被严格的 使用小鼠和斑马鱼细胞，通过生物化学和最先进的成像相结合进行分析 和组织，包括我们拥有专业知识的运动神经元。如果实现的话，我们的目标将会发现新颖的 SMN 结合伴侣和 SMNTud 在生物分子缩合中的功能。监管潜力和 DMA 修饰的动态性（组织特异性和疾病病因学的来源）将被确定。 DMA-SMNTud 相互作用模块的鉴定将为 SMA 提供新的治疗方法。