Nanoscopic Membrane Modulations Induced by Nanoscale Oligomers

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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