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Research for the role of dystrophic neurites in neuronal cell dysfunction of Polyglutamine disease

营养不良性神经突在多聚谷氨酰胺病神经细胞功能障碍中的作用研究

基本信息

批准号：
13670635
负责人：
ONODERA Osamu
金额：
$ 2.3万
依托单位：
Niigata University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2001
资助国家：
日本
起止时间：
2001 至 2002
项目状态：
已结题

项目摘要

To understand the pathogenetic mechanisms underlying polyglutamine diseases, we investigated the mechanisms of the formation of aggregate bodies containing expanded polyQ stretches, focusing on dentatorubral-pallidoluysian atrophy (DRPLA).We demonstrated that the expression of a truncated DRPLA protein containing expanded polyQ stretches in COS-7 cells resulted in the formation of perinuclear aggregate bodies that are co-localized with γ-tubulin, a protein marker for the microtubules-organizing center (MTOC). A collapsed vimentin network surrounded these aggregate bodies. Furthermore, disruption of the microtubules with nocodazole resulted in the formation of small aggregate bodies that were scattered throughout the cytoplasm. These findings suggest that the truncated DRPLA proteins containing expanded polyQ stretches unfold and form small aggregate bodies in the cell periphery. These aggregates move on microtubules to the MTOC, where they remain as distinct "aggresomes".Then we have established several stable cell lines, which expressed polyQ stretches under control of tetracycline. In these cell lines, the integration sites are identical so we could exclude the effect of a chromosomal localization of integrated cDNA. Using these stable cell lines we have seen the turnover rates of polyQ stretch is different.In future we will analysis the effects of the difference of the fate of polyQ stretch by their length on neuronal cell function, specially their axonal transport system.

为了了解多聚谷氨酰胺疾病的发病机制，我们研究了含有扩展polyQ片段的聚集体的形成机制，重点是齿状核红核-苍白球路易氏体萎缩（DRPLA）。我们证明，在COS-7细胞中表达含有扩展polyQ片段的截短DRPLA蛋白导致核周聚集体的形成，该聚集体与γ-微管蛋白共定位，微管组织中心（MTOC）的蛋白质标记物。一个塌陷的波形蛋白网络包围着这些聚集体。此外，用诺考达唑破坏微管导致分散在整个细胞质中的小聚集体的形成。这些发现表明，截短的DRPLA蛋白含有扩展polyQ伸展展开，并在细胞外周形成小的聚集体。这些聚集体在微管上移动到MTOC，在那里它们保持为不同的“攻击体”。然后，我们建立了几个稳定的细胞系，它们在四环素的控制下表达polyQ片段。在这些细胞系中，整合位点是相同的，因此我们可以排除整合的cDNA的染色体定位的影响。利用这些稳定的细胞系，我们已经看到polyQ拉伸的周转率是不同的。在未来，我们将分析polyQ拉伸的命运的差异，其长度对神经元细胞功能的影响，特别是其轴突运输系统。