Nanoscopic Membrane Modulations Induced by Nanoscale Oligomers

亨廷顿病（HD）是由突变亨廷顿蛋白（Htt）引起的破坏性神经元疾病， 多聚谷氨酰胺（polyQ）束的异常扩张。突变型Htt具有形成 预纤维状和纤维状聚集体。尽管聚集体决定的毒性已被广泛建议用于 过多的神经元疾病，淀粉样蛋白如何导致HD中的中等多刺神经元死亡的机制 仍不清楚虽然纤维状聚集体通常存在于斑和内含体中，但小的 现在认为寡聚物是许多神经变性疾病的主要分子毒素。 基于寡聚体假说和膜组件在淀粉样蛋白毒性中的明显作用，我们 假设突变型Htt纳米级寡聚体负责调节膜特性， 导致细胞功能的进行性丧失，例如钙离子的不受调节的流动和异常的形状 突触和线粒体膜的转化。我们进一步假设， 相互作用是脂质特异性的。由于脂质谱是细胞依赖性和年龄依赖性的，脂质特异性可能 作为一个合理的原因，可变的细胞脆弱性相同的淀粉样物质。虽然polyQ- 膜相互作用已被广泛研究，预原纤和原纤聚集体的混合物通常是 礼物因此，很难区分哪种聚集物种是造成这种现象的原因。 观察到对膜性能的影响。我们将首先从原纤维中分离可溶性polyQ寡聚体， 单体。然后，我们将使用实验方法来研究寡聚体膜相互作用。在Aim中 1，我们将寻求确定纳米级低聚物对纳米材料的微观和纳米级性质的影响。 脂膜基于特定脂质在HD发病机制中的意义，我们将制备脂质 具有确定的脂质组成的膜。我们的脂质依赖性研究将阐明特定脂质的作用 在控制低聚物毒性方面。在目标2中，我们将进行拉曼光谱实验来研究 脂质膜的原子水平变化。拉曼光谱数据将揭示低聚物如何干扰脂质i） 链内构象顺序和ii）链间堆积顺序。生物物理工具的组合将 在这个项目中，包括高分辨率原子力显微镜（AFM），基于AFM的力 光谱法、囊泡渗漏测定法、微量吸液管抽吸法和拉曼光谱法。生物物理 在这个项目中阐述的方法可以转移到一系列的低聚物形成的调查， 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， 阐明寡聚体控制的毒性的分子基础。 1/1

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