Nanoscopic Membrane Modulations Induced by Nanoscale Oligomers

纳米级低聚物诱导的纳米膜调节

• 批准号: 10790511
• 负责人: Jianjun Pan
• 金额: $ 14.17万
• 依托单位: UNIVERSITY OF SOUTH FLORIDA
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-09-08 至 2025-08-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10790511
• 关键词: American; Amyloid; Atomic Force Microscopy; Biological Assay; Calcium ion; Cardiolipins; Cell physiology; Cells; Cholesterol; Cognitive; Data; Disease; Elasticity; Extravasation; FDA approved; Fingerprint; Future; Ganglioside GM1; Genetic Diseases; Goals; Huntington Disease; Huntington gene; Inclusion Bodies; Investigation; Lateral; Lipid Bilayers; Lipids; Liquid substance; Mechanics; Mediating; Membrane; Membrane Lipids; Microscopic; Molecular; Molecular Conformation; Morphology; Motor; Neurodegenerative Disorders; Neurons; Pathogenesis; Penetration; Peptides; Permeability; Pharmaceutical Preparations; Pilot Projects; Property; Proteins; Raman Spectrum Analysis; Research; Resolution; Risk; Role; Shapes; Specificity; Spectrum Analysis; Stretching; Structure; Symptoms; Synaptic Membranes; Toxic effect; Toxin; Vertebral column; Vesicle; age related; aspirate; base; biophysical techniques; biophysical tools; experimental study; lipid I; membrane assembly; misfolded protein; mitochondrial membrane; monomer; mutant; nano; nanomechanics; nanoscale; neuron loss; polyglutamine; protein aggregation; release of sequestered calcium ion into cytoplasm; unilamellar vesicle

Huntington’s disease (HD) is a devastating neuronal disorder caused by the mutant huntingtin (Htt) protein with an abnormal expansion of the polyglutamine (polyQ) tract. Mutant Htt has an intrinsic propensity of forming prefibrillar and fibrillar aggregates. Although aggregates-dictated toxicity has been widely suggested for a plethora of neuronal disorders, the mechanism of how amyloids cause medium spiny neuronal death in HD remains unclear. Although fibrillar aggregates are commonly found in plagues and inclusions bodies, small oligomers are now considered as the dominant molecular toxins underlying many neurodegenerative diseases. Based on the oligomer hypothesis and the apparent role of membrane assemblies in amyloid toxicity, we hypothesize that nanoscale oligomers of mutant Htt are responsible for modulating membrane properties, leading to progressive loss of cellular functions such as unregulated flux of calcium ions and abnormal shape transformation of synaptic and mitochondrial membranes. We further hypothesize that oligomer-membrane interactions are lipid specific. Because lipid profile is cell dependent and age dependent, lipid specificity could serve as a plausible cause for variable cell vulnerability to the same amyloid species. Although polyQ- membrane interactions have been widely studied, a mixture of prefibrillar and fibrillar aggregates are often present. Consequently, it is challenging to distinguish which aggregated species is responsible for the observed effects on membrane properties. We will first separate soluble polyQ oligomers from fibrils and monomers. We will then use experimental approaches to investigate oligomer-membrane interactions. In Aim 1, we will seek to determine the effects of nanoscale oligomers on microscopic and nanoscopic properties of lipid membranes. Based on the significance of specific lipids in HD pathogenesis, we will prepare lipid membranes with defined lipid compositions. Our lipid-dependent studies will elucidate the role of specific lipids in governing oligomer toxicity. In Aim 2, we will conduct Raman spectroscopy experiments to investigate atomic-level changes of lipid membranes. Raman spectral data will unveil how oligomers perturb the lipid i) intra-chain conformational order and ii) inter-chain packing order. A combination of biophysical tools will be employed in this project, including high-resolution atomic force microscopy (AFM), AFM-based force spectroscopy, vesicle leakage assay, micropipette aspiration, and Raman spectroscopy. The biophysical approaches elaborated in this project can to transferred to investigations of an array of oligomers formed by different amyloidogenic proteins. Therefore, our research will pave the way for future studies aimed at elucidating molecular bases of oligomer-governed toxicity. 1/1

亨廷顿氏病（HD）是由突变亨廷顿蛋白（HTT）蛋白引起的毁灭性神经元疾病 聚谷氨酰胺（Polyq）裂纹的异常膨胀。突变htt具有形成的内在诺言 预纤维和纤维骨料。 过多的神经元疾病，淀粉样蛋白如何导致培养基神经元死亡的机制 尚不清楚。尽管原纤维骨料通常在鼠疫和包裹体体中发现，但很小 现在，低聚物被认为是许多神经退行性疾病的主要分子毒素。 基于低聚物假说和膜组件在淀粉样蛋白毒性中的明显作用，我们 假设突变体HTT的纳米级低聚物负责调节膜特性， 导致细胞功能的逐渐丧失，例如钙离子不受管制的通量和异常形状 突触和线粒体膜的转化。我们进一步假设低聚物膜 相互作用是脂质的。由于脂质谱是细胞依赖性和年龄依赖性的，因此脂质特异性可以 可作为可变细胞脆弱性与同一淀粉样物种的合理原因。虽然polyq- 膜相互作用已被广泛研究，前纤维和纤维骨料的混合物通常是 展示。因此，区分哪些汇总物种负责是挑战 观察到对膜特性的影响。我们将首先将固体polyq低聚物与原纤维分开 单体。然后，我们将使用实验方法研究寡聚物 - 膜相互作用。目标 1，我们将寻求确定纳米级低聚物对显微镜和纳米镜的影响的影响 脂质膜。基于特定脂质在HD发病机理中的重要性，我们将制备脂质 具有定义的脂质组成的膜。我们的脂质依赖性研究将阐明特定脂质的作用 在AIM 2中，我们将进行拉曼光谱实验以研究 脂质膜的原子水平变化。拉曼光谱数据将揭示低聚物如何扰动脂质i） 链内构象顺序和ii）链间填料顺序。生物物理工具的结合将是 在该项目中使用，包括高分辨率原子力显微镜（AFM），基于AFM的力 光谱，囊泡泄漏测定，微孔抽吸和拉曼光谱。生物物理 该项目中详细阐述的方法可以转移到对由 不同的淀粉样蛋白。因此，我们的研究将为未来的研究铺平道路 阐明寡聚物毒性的分子碱基。 1/1