Poly(ADP-ribose)-dependent TDP-43 pathology in oxidative stress (R21)

氧化应激中聚 (ADP-核糖) 依赖性 TDP-43 病理学 (R21)

• 批准号: 10753095
• 负责人: Shaida A. Andrabi
• 金额: $ 40.84万
• 依托单位: UNIVERSITY OF ALABAMA AT BIRMINGHAM
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-09-01 至 2025-08-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10753095
• 关键词: Acceleration; Address; Affect; Alzheimer' s Disease; Alzheimer' s disease related dementia; Amyotrophic Lateral Sclerosis; Area; Binding; Biochemical; Biological; Biological Assay; Cell Death; Cell Nucleus; Cells; Cessation of life; Cryoelectron Microscopy; Cytoplasm; Cytosol; DNA-Binding Proteins; Data; Defect; Dementia; Deuterium; Disease; Event; Exposure to; Frontotemporal Dementia; Hydrogen; Hydrogen Peroxide; Imaging Techniques; Impairment; Inclusion Bodies; Length; Ligation; Limbic System; Mass Spectrum Analysis; Mediating; Modeling; Molecular Conformation; Mus; Mutation; Nature; Neurodegenerative Disorders; Neurons; Nuclear; Nuclear Export; Nuclear Import; Nuclear Localization Signal; Nuclear Protein; Oxidative Stress; Oxidative Stress Induction; Pathologic; Pathologic Processes; Pathology; Pathway interactions; Physiological; Play; Poly Adenosine Diphosphate Ribose; Poly(ADP-ribose) Polymerase Inhibitor; Poly(ADP-ribose) Polymerases; Polymers; Property; Protein Export Pathway; Protein translocation; Proteins; Publishing; RNA Recognition Motif; Role; Signal Transduction; Signaling Molecule; Solubility; Stress; Structure; Temporal Lobe; Time; Toxic effect; Transactivation; Variant; biophysical properties; confocal imaging; conformational alteration; disease-causing mutation; exportin 1 protein; inhibitor; nervous system disorder; neuron loss; protein TDP-43; response; virtual

Abstract Frontotemporal dementia (FTD) is an Alzheimer-related dementia disease (ADRD) and involves nuclear egress and cytoplasmic Transactivation response DNA binding protein 43 (TDP-43). The mechanisms for pathological export of TDP-43 or its accumulation in the cytoplasm is not clearly defined. Although, studies on disease-causing mutations have revealed that defects in nuclear import may be in part responsible for TDP-43 accumulation in the cytosol as these mutations change the properties of the protein. However, it remains largely unclear how TDP-43 accumulates in the cytosol, as the majority of the TDP-43 dependent pathologies are sporadic. TDP-43 contains two RNA recognition motifs (RRMs), a nuclear export signal (NES), and a nuclear localization signal (NLS) in its N-terminus, and binds poly (ADP-ribose) polymer (PAR) via a PAR-binding motif embedded in its NLS. Studies have shown that PAR inhibitors can inhibit TDP-43 pathology, and binding of PAR to TDP-43 can change its biophysical characteristics in solution. Exportin 1 (XPO1) mediates the NES-dependent export of proteins from the nucleus. TDP-43 is a 43kDa protein that may not require XPO1-dependent export under basal conditions but our data indicates that under neuronal stress conditions TDP-43 egresses the nucleus via XPO-1 dependent export. We hypothesize that in disease conditions like FTD type ADRD and related diseases; PAR binds TDP-43 in the nucleus and facilitates its interaction with XPO-1, which subsequently enhances the egress of TDP-43 from the nucleus to cytosol. This proposal is supported our preliminary data that both PARP inhibitor BMN673 and XPO-1 inhibitor KPT- 185 inhibit the cytosolic accumulation of TDP-43 in neurons and TDP-43 and XPO1 interact following PARP activation. In this proposal, we plan to study the alterations in TDP-43 in mouse cortical neurons exposed to oxidative stress. Oxidative stress is a common pathological process mediating protein mislocalization, aggregation, and cell death in virtually all neurological disorders including FTD. Oxidative stress via H2O2 in cortical neurons mediates a robust TDP-43 cytosolic localization. Therefore, it is likely that pathophysiological mechanisms identified in the oxidative stress model in cortical neurons can closely overlap/mimic pathological mechanisms in FTD and other ADRD involving changes/mislocalization of TDP-43. We will use biochemical, cell biological and imaging techniques in combination with proximity ligation assays, CryoEM, and Hydrogen/Deuterium Exchange mass spectrometry to assess the PAR-dependent alteration in TDP-43 and its interaction with XPO-1 that leads to an increase in the release of TDP-43 from the nucleus and subsequent accumulation/aggregation in the cytoplasm. These studies will help in understanding the pathophysiological mechanisms of TDP-43 accumulation in FTD, ALS, and other ADRD/dementia related diseases involving TDP-43

抽象的 额颞叶痴呆 (FTD) 是一种阿尔茨海默相关痴呆病 (ADRD)，涉及核 出口和细胞质反式激活反应 DNA 结合蛋白 43 (TDP-43)。的机制 TDP-43 的病理输出或其在细胞质中的积累尚不清楚。虽然，研究 对引起疾病的突变的研究表明，核输入缺陷可能是导致疾病的部分原因 由于这些突变改变了蛋白质的特性，TDP-43 在细胞质中积累。然而，它 目前仍不清楚 TDP-43 如何在细胞质中积累，因为大多数 TDP-43 依赖 病理是散发的。 TDP-43 包含两个 RNA 识别基序 (RRM)，一种核输出信号 (NES)，其 N 末端有核定位信号 (NLS)，并结合聚 (ADP-核糖) 聚合物 (PAR) 通过嵌入其 NLS 中的 PAR 结合基序。研究表明PAR抑制剂可以抑制TDP-43 病理学上，PAR 与 TDP-43 的结合可以改变其在溶液中的生物物理特性。出口1 (XPO1) 介导 NES 依赖性蛋白质从细胞核的输出。 TDP-43 是一种 43kDa 的蛋白质， 在基础条件下可能不需要 XPO1 依赖性输出，但我们的数据表明在神经元条件下 应激条件下，TDP-43 通过 XPO-1 依赖的输出从细胞核中流出。我们假设在疾病中 FTD 型 ADRD 等病症及相关疾病； PAR 与细胞核中的 TDP-43 结合并促进其 与 XPO-1 相互作用，随后增强 TDP-43 从细胞核到细胞质的流出。 该提议得到了我们的初步数据的支持，即 PARP 抑制剂 BMN673 和 XPO-1 抑制剂 KPT- 185 抑制神经元中 TDP-43 的胞质积聚，并且 TDP-43 和 XPO1 在 PARP 后相互作用 激活。在本提案中，我们计划研究暴露于 氧化应激。氧化应激是介导蛋白质错误定位的常见病理过程， 几乎所有神经系统疾病（包括 FTD）中的聚集和细胞死亡。通过 H2O2 产生的氧化应激 皮质神经元介导强大的 TDP-43 胞质定位。因此，病理生理学上可能是 皮层神经元氧化应激模型中发现的机制可以紧密重叠/模仿病理学 FTD 和其他 ADRD 中涉及 TDP-43 变化/错误定位的机制。 我们将使用生物化学、细胞生物学和成像技术与邻近连接测定相结合， CryoEM 和氢/氘交换质谱法可评估 PAR 依赖性变化 TDP-43 及其与 XPO-1 的相互作用导致 TDP-43 从细胞核中释放增加 以及随后在细胞质中的积累/聚集。这些研究将有助于理解 FTD、ALS 和其他 ADRD/痴呆相关的 TDP-43 积累的病理生理机制 涉及 TDP-43 的疾病