Ultrastructure of nuclear aggregates formed by expressing an expanded polyglutamine
Ultrastructure of nuclear aggregates formed by expressing an expanded polyglutamine
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DOI：
10.1016/s0006-291x(02)00498-9
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发表时间：
2002-06-07
影响因子：
3.1
通讯作者：
Kanazawa, I
中科院分区：
文献类型：
--
作者：
Hazeki, N;Tsukamoto, T;Kanazawa, I
在亨廷顿病(HD)患者的大脑中观察到了核内包涵体。瞬时表达HD外显子1的细胞含有74个谷氨酰胺重复序列，这些重复序列与绿色荧光蛋白(GFP)和核定位序列(NLS)相连，在细胞核中含有聚集体。通过离心分级和荧光激活细胞分选(FACS)对聚集体进行纯化。在SDS样品缓冲液中对集合体进行热处理，导致致密的集合体核心消失，并生成由原纤维组成的篮状结构。生物化学分析表明，HD外显子1-GFP融合蛋白是主要成分。异质性核糖核蛋白F和H、组蛋白和泛素被发现与聚集体有关。我们的观察表明，Huntingtin的N-末端片段可能组织了聚集体的骨架结构，并可能通过捕获聚集体中的其他蛋白质来干扰正常的细胞功能。(C)2002年埃尔塞维尔科学公司(美国)。版权所有。
Intranuclear inclusions have been observed in the brains of patients affected with Huntington's disease (HD). Neuro 2A cells that transiently expressed HD exon 1 bearing 74 glutamine repeats linked to the green fluorescent protein (GFP) and the nuclear localization sequence (NLS) contained aggregates in nuclei. The aggregates were purified by fractionation with centrifugation followed by fluorescence-activated cell sorting (FACS). Heat treatment of the aggregate in an SDS sample buffer caused the dense aggregate cores to disappear and generated a basket-like structure composed of fibrils. Biochemical analysis of the aggregates revealed that the HD exon 1-GFP fusion protein was the major component. The heterogeneous nuclear ribonucleoproteins F and H, histones and ubiquitin were found to be associated with the aggregates. Our observations suggest that the N-terminal fragment of huntingtin may organize the skeletal structure of the aggregates and may disturb normal cellular functions by trapping other proteins within the aggregates. (C) 2002 Elsevier Science (USA). All rights reserved.