Onodera O, Burke J R, Miller S E, Hester S, Tsuji S, Roses A D, Strittmatter W J
Department of Medicine (Neurology), Duke University Medical Center, Durham, North Carolina 27710, USA.
Biochem Biophys Res Commun. 1997 Sep 18;238(2):599-605. doi: 10.1006/bbrc.1997.7337.
Six inherited neurologic diseases, including Huntington's disease, result from the expansion of a CAG domain of the disease genes to produce a domain of more than 40 glutamines in the expressed protein. The mechanism by which expansion of this polyglutamine domain causes disease is unknown. Recent studies demonstrated oligomerization of polyglutamine-domain proteins in mammalian neurons. To study oligomerization of polyglutamine proteins and to identify heterologous protein interactions, varying length polyglutamine-green fluorescent protein fusion proteins were expressed in cultured COS-7 cells. The 19- and 35-glutamine fusion proteins (non-pathologic length) distributed diffusely throughout the cytoplasm. In contrast, 56- and 80-glutamine fusion proteins (pathologic length) formed fibrillar arrays resembling those previously observed in neurons in Huntington's disease and in a transgenic mouse model. These aggregates were intranuclear and intracytoplasmic. Intracytoplasmic aggregates were surrounded by collapsed intermediate filaments. The intermediate filament protein vimentin co-immunoisolated with expanded polyglutamine fusion proteins. This cellular model will expedite investigations into oligomerization of polyglutamine proteins and their interactions with other proteins.
包括亨廷顿舞蹈症在内的六种遗传性神经疾病，是由于疾病基因的CAG结构域发生扩增，从而在表达的蛋白质中产生一个含有超过40个谷氨酰胺的结构域所致。这种多聚谷氨酰胺结构域的扩增导致疾病的机制尚不清楚。最近的研究表明，多聚谷氨酰胺结构域蛋白在哺乳动物神经元中会发生寡聚化。为了研究多聚谷氨酰胺蛋白的寡聚化并鉴定异源蛋白相互作用，在培养的COS-7细胞中表达了不同长度的多聚谷氨酰胺-绿色荧光蛋白融合蛋白。含有19个和35个谷氨酰胺的融合蛋白（非致病长度）在整个细胞质中呈弥散分布。相比之下，含有56个和80个谷氨酰胺的融合蛋白（致病长度）形成了纤维状阵列，类似于先前在亨廷顿舞蹈症患者的神经元以及转基因小鼠模型中观察到的情况。这些聚集体存在于细胞核内和细胞质内。细胞质内的聚集体被塌陷的中间丝所包围。中间丝蛋白波形蛋白与扩增的多聚谷氨酰胺融合蛋白进行了共免疫分离。这个细胞模型将加快对多聚谷氨酰胺蛋白寡聚化及其与其他蛋白相互作用的研究。
