Pathological aggregation of TDP-43 in age-related neurodegenerative diseases

TDP-43 在年龄相关神经退行性疾病中的病理聚集

基本信息

批准号：
9812745
负责人：
William Michael Babinchak
金额：
$ 4.95万
依托单位：
CASE WESTERN RESERVE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2018
资助国家：
美国
起止时间：
2018-09-14 至 2021-05-13
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9812745
关键词：
Adopted Aging Alzheimer&apos s Disease Amino Acids Amyloid Amyloid Fibrils Amyotrophic Lateral Sclerosis Atomic Force Microscopy Brain C-terminal Caspase Cerebrum Clinical Cystic Fibrosis Transmembrane Conductance Regulator Cytoplasmic Granules DNA Binding DNA-Binding Proteins Data Deposition Development Disease Electron Spin Resonance Spectroscopy Fiber Frontotemporal Dementia Frontotemporal Lobar Degenerations Genetic Polymorphism Immobilization Individual Knowledge Laboratories Liquid substance Mediating Messenger RNA Modeling Molecular Morphology Nerve Degeneration Neuraxis Neurodegenerative Disorders Pathologic Patients Periodicity Phase Phase Transition Phenotype Production Protein Fragment Proteins RNA RNA Splicing RNA-Binding Proteins Reporting Role Sclerosis Seeds Site Spin Labels Structural Models Structure System Testing Therapeutic Toxic effect Transact Transcript Ubiquitin Water X ray diffraction analysis age related age related neurodegeneration aging population amyloid fibril formation base experimental study hippocampal sclerosis insight lens nucleocytoplasmic transport polymerization prion-like protein aggregate protein aggregation protein protein interaction recruit response stress granule

项目摘要

The transactive response DNA-binding protein of 43kDa (TDP-43) is a DNA- and RNA-binding protein recognized to form pathological amyloid inclusions in the brains of patients with age-related neurodegenerative disorders, including Alzheimer’s disease, frontotemporal lobar degeneration, and cerebral age-related TDP-43 with sclerosis. Interestingly, these inclusions have also been found in subsets of clinically asymptomatic patients of the aging population, highlighting the need to understand the structural basis of disease development. Current evidence suggests that TDP-43 aggregation is facilitated by the production of C-terminal fragments of the protein that have a high propensity to form amyloid fibrils due to the presence of a low complexity domain (LCD)—a region with low amino acid diversity. In addition to forming amyloid fibrils, the TDP-43 LCD was recently identified to undergo liquid-liquid phase separation (LLPS), a phenomenon whereby protein-protein interactions induce dynamic, reversible coacervation. Prolonged or repetitive coacervation is thought to drive pathological deposition of aggregates; however, a direct connection between TDP-43 LLPS and aggregation that would provide a mechanistic basis for Alzheimer’s disease and related neurodegenerative disorders has not been established. To better understand the structural underpinnings of TDP-43 amyloid aggregates and the role of LLPS in their formation, LCD aggregation conditions were screened and LLPS was identified to regulate amyloid polymerization. Because a comprehensive structural model for TDP-43 fibrils has yet to be proposed, electron paramagnetic resonance (EPR) spectroscopy was subsequently utilized to identify immobilized residues of the LCD that likely constitute the fibrillar core, the structural building block of amyloids. Interestingly, EPR spectra of residues within the fibrillar core did not suggest that the TDP-43 LCD adopts the most commonly observed cross-β organization—a parallel, in-register organization of β-strands that is observed among other protein aggregates in Alzheimer’s disease. As such, we hypothesized that TDP-43 fibrils adopt a non-canonical structure. To test this hypothesis, we first propose a comprehensive EPR-based approach to (i) identify the full extent of residues involved in the fibrillar core, ascertain which residues participate in β-strands and (ii) identify whether a parallel, in-register motif is present. If this motif can be ruled out, (iii) fiber X-ray diffraction will be used to differentiate non-canonical structures (antiparallel, β-solenoid), and (iv) EPR-derived spin-spin distances will be used to determine the organization of residues at the intermolecular interface between fibrillar subunits. Preliminary data further suggest that LLPS might facilitate formation of an extended fibrillar core. Our second aim therefore will identify whether LLPS alters the underlying cross-β structure of TDP-43 amyloids and propagates a structural polymorphism. This approach will thus provide structural insights about TDP-43 amyloids and ascertain whether LLPS might facilitate differential aggregation of amyloid fibrils in Alzheimer’s disease and related neurodegenerative disorders.

The transactive response DNA-binding protein of 43kDa (TDP-43) is a DNA- and RNA-binding protein recognized to form pathological amyloid inclusions in the brains of patients with age-related neurodegenerative disorders, including Alzheimer’s disease, frontotemporal lobar degeneration, and cerebral age-related TDP-43 with sclerosis.有趣的是，这些夹杂物也在老龄化人群的临床不对称患者的子集中发现，强调了了解疾病发展的结构性基础的必要性。当前的证据表明，由于存在低复杂性结构域（LCD）（一个具有低氨基酸多样性的区域），该蛋白质的C末端片段的产生很高，因此TDP-43聚集得到了形成淀粉样蛋白原纤维的高希望。除了形成淀粉样蛋白原纤维外，最近还确定了TDP-43 LCD进行液态液相分离（LLP），这是一种蛋白质 - 蛋白质相互作用的现象，蛋白质 - 蛋白质相互作用诱导了动态，可逆的关联。人们认为延长或重复的关联可以驱动聚集体的病理沉积。然而，尚未建立TDP-43 LLP与聚集的直接联系，这将为阿尔茨海默氏病和相关的神经退行性疾病提供机械基础。为了更好地了解TDP-43淀粉样蛋白聚集体的结构基础和LLP在其形成中的作用，筛选了LCD聚集条件，并确定了LLPS以调节淀粉样蛋白聚合。由于尚未提出了TDP-43原纤维的综合结构模型，因此随后利用电子顺磁共振（EPR）光谱法来识别可能构成纤维核心核心核心的固定的LCD的保留率，该保留可能构成纤维核的结构构建块。有趣的是，原纤维芯内残留物的EPR光谱并未表明TDP-43 LCD采用最常见的跨β组织，这是一个平行的β链组织，在阿尔茨海默氏病中其他蛋白质聚集体中观察到的β链。因此，我们假设TDP-43原纤维采用非规范结构。为了检验这一假设，我们首先提出了一种基于EPR的综合方法，以（i）确定纤维芯中涉及的残留物的全部程度，确定哪些保留参与β链，以及（ii）确定是否存在平行的，内置的基序。如果可以排除该基序，（iii）（iii）纤维X射线衍射将用于区分非经典结构（反平行，β-甲素），并且（IV）EPR衍生的自旋旋转距离将用于确定纤维中纤维中界面界面界面界面界面界面的残基的组织。进一步的数据进一步表明，LLP可能会准备扩展的纤维芯的形成。因此，我们的第二个目标将确定LLP是否会改变TDP-43淀粉样蛋白的基本交叉结构并传播结构性多态性。因此，这种方法将提供有关TDP-43淀粉样蛋白的结构见解，并确定LLP是否可以促进阿尔茨海默氏病和相关神经退行性疾病中淀粉样蛋白原纤维的差异聚集。