ROLE OF CHAIN LENGTH AND SEQUENCE CONTEXTS ON POLYGLUTAMINE OLIGOMERIZATION

 描述（由申请人提供）：亨廷顿病（HD）是一种破坏性神经退行性疾病，由亨廷顿（Htt）基因外显子1中CAG密码子的突变扩增引起。CAG重复扩增导致亨廷顿蛋白的N-末端区域中的多聚谷氨酰胺结构域扩增。Htt的N-末端片段（Htt-NTF）通过全长蛋白质的蛋白水解或通过突变mRNA转录物的异常剪接产生。这些碎片最终成为不溶性的核内包涵体在中型棘和小脑神经元。Htt-NTF包含17个残基的N-末端片段（N17）、polyQ结构域和38个残基的富含脯氨酸的C-末端片段（C38）。我们已经表明，N17拉伸加速原纤维的形成，而C38拉伸不稳定的不溶性原纤维和稳定大的可溶性球形聚集体。最近在转基因小鼠中的研究表明，N17的缺失导致极端的神经毒性和快速的神经功能下降。这些特征代表HD的发作和进展。体内研究与我们最近的发现一起，导致的建议，可溶性球形聚集体，这是稳定的C38，可能是HD的神经毒性的来源。在这种竞争性的更新应用中，我们将采用一种独特的方法，将新颖的多尺度计算机模拟与体外生物物理实验相结合，以提高我们对Htt-NTFs的聚集机制和相行为的理解。我们的研究是由我们的粗粒度模拟的新方法的帮助。这些模拟结合新的体外实验将帮助我们了解C38模块如何稳定可溶性球形聚集体，同时减缓缺乏N17模块的Htt-NTFs中从可溶性球体到不溶性原纤维的转化。我们还试图了解N17模块的原纤维稳定作用如何与C38在外显子1中的原纤维去稳定作用竞争，所述C38跨越具有不同长度的polyQ束的Htt-NTFs。清楚地理解由于polyQ长度、侧翼序列模块N17和C38之间的相互作用而形成的聚集机制和结构对于辨别Htt-NTF聚集和神经毒性之间的联系是绝对必要的。

 DESCRIPTION (provided by applicant): Huntington's disease (HD) is a devastating neurodegenerative disease that is caused by mutational expansion of the CAG codon in exon 1 of the huntingtin (Htt) gene. The CAG repeat expansion leads to an expanded polyglutamine domain in the N-terminal region of the huntingtin protein. N-terminal fragments of Htt (Htt-NTFs) are generated by proteolysis of the full-length protein or by aberrant splicing of mutant mRNA transcripts. These fragments end up as insoluble intranuclear inclusions within medium spiny and cerebellar neurons. Htt-NTFs encompass a 17-residue N-terminal stretch (N17), a polyQ domain, and a 38-residue proline-rich C-terminal stretch (C38). We have shown that the N17 stretch accelerates fibril formation whereas the C38 stretch destabilizes insoluble fibrils and stabilizes large soluble spherical aggregates. Recent studies in transgenic mice show that the deletion of N17 leads to extreme neurotoxicity and rapid neurological decline. These features typify the onset and progression of HD. The in vivo studies taken together with our recent findings, lead to the proposal that soluble spherical aggregates, which are stabilized by C38, might be the source of neurotoxicity in HD. In this competitive renewal application, we will pursue a unique approach that combines novel multiscale computer simulations with in vitro biophysical experiments to advance our understanding of the aggregation mechanisms and phase behavior of Htt-NTFs. Our studies are aided by our new methodologies for coarse-grained simulations. These simulations combined with novel in vitro experiments will help us understand how the C38 module stabilizes soluble spherical aggregates while slowing the conversion from soluble spheres to insoluble fibrils in Htt-NTFs that lack the N17 module. We also seek to understand how the fibril stabilizing effect of the N17 module competes with the fibril destabilizing effects of C38 in exon 1 spanning Htt-NTFs with polyQ tracts of different lengths. A clear understanding of the aggregation mechanisms and the structures that form as the result of the interplay among polyQ length, the flanking sequence modules N17 and C38 is absolutely necessary to discern the connection between Htt-NTF aggregation and neurotoxicity.