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

亨廷顿病 (HD) 是一种毁灭性的神经系统疾病，由突变亨廷顿 (Htt) 蛋白引起， 突变体 Htt 的异常扩张具有形成的内在倾向。 尽管聚集体决定的毒性已被广泛认为是前原纤维和原纤维聚集体。 过多的神经元疾病，淀粉样蛋白如何导致 HD 中型多刺神经元死亡的机制 尽管纤维状聚集体常见于斑块和包涵体中，但仍不清楚。 寡聚物现在被认为是许多神经退行性疾病的主要分子毒素。 基于寡聚物假说和膜组件在淀粉样蛋白毒性中的明显作用，我们 研究发现突变体 Htt 的纳米级寡聚物负责调节膜特性， 导致细胞功能逐渐丧失，例如钙离子流量不受控制和形状异常 我们进一步捕获了突触和线粒体膜的转化。 由于脂质谱具有细胞依赖性和年龄依赖性，因此脂质特异性可能存在。 作为不同细胞对同一淀粉样蛋白种类易感性的合理原因。 膜相互作用已被广泛研究，通常是前原纤维和原纤维聚集体的混合物 目前，很难区分是哪种聚集物种造成的。 我们首先将可溶性 PolyQ 低聚物与原纤维分离并观察对膜性能的影响。 然后我们将使用实验方法来研究低聚物-膜相互作用。 1，我们将寻求确定纳米级低聚物对微观和纳米性质的影响 基于特定脂质在HD发病机制中的重要性，我们将制备脂质。 我们的脂质依赖性研究将阐明特定脂质的作用。 在目标 2 中，我们将进行拉曼光谱实验来研究。 脂质膜的原子级变化将揭示低聚物如何扰乱脂质 i) 链内构象顺序和 ii) 链间包装顺序将是生物物理工具的组合。 该项目中采用的技术包括高分辨率原子力显微镜 (AFM)、基于 AFM 的力 光谱学、囊泡渗漏测定、微量移液器抽吸和拉曼光谱学。 该项目中阐述的方法可以转移到对由以下物质形成的一系列低聚物的研究 因此，我们的研究将为未来的研究铺平道路。 阐明低聚物控制毒性的分子基础。 1/1