HspB8, a small heat shock protein mutated in human neuromuscular disorders, has in vivo chaperone activity in cultured cells
HspB8, a small heat shock protein mutated in human neuromuscular disorders, has in vivo chaperone activity in cultured cells
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DOI：
10.1093/hmg/ddi174
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发表时间：
2005-06-15
影响因子：
3.5
通讯作者：
Landry, J
中科院分区：
文献类型：
--
作者：
Carra, S;Sivilotti, M;Landry, J
小热休克蛋白（sHsp）家族在哺乳动物中由10个成员组成，其中4个在与蛋白质聚集体积累相关的疾病中被发现发生突变。尽管许多sHsp在体外无细胞条件下表现出分子伴侣活性，但它们在正常细胞环境下的体内活性尚不清楚。在本研究中，我们以聚谷氨酰胺蛋白Htt43Q为模型，研究了sHsp HspB8/Hsp22在细胞中阻止蛋白质聚集的能力。在对照条件下，Htt43Q在由sds不溶性聚集体组成的核周包裹体中积累。与Htt43Q共转染后，HspB8偶尔会被困在内含物中；然而，在大多数细胞中，HspB8阻断包涵体的形成。生化分析表明，HspB8与阳性对照Hsp40一样有效地抑制了sds不溶性Htt43Q的积累。然后，在蛋白酶体和自噬抑制剂阻断蛋白质降解的情况下，Htt43Q在sds可溶部分中积累。而另一种sHsp Hsp27/HspB1和α B-crystallin/HspB5则无影响。这表明HspB8作为分子伴侣，维持Htt43Q处于可快速降解的可溶性状态。对Hsp27-HspB8嵌合蛋白的分析表明，HspB8的c端结构域包含伴侣蛋白活性所需的特定序列。在人类运动神经病变中发现的赖氨酸141区域的错义突变显著降低了该蛋白的伴侣活性。因此，HspB8伴侣活性的降低可能有助于这些疾病的发展。
The family of small heat shock proteins (sHsp) is composed of 10 members in mammals, four of which are found mutated in diseases associated with the accumulation of protein aggregates. Though many sHsp have demonstrated molecular chaperone activity in vitro in cell-free conditions, their activity in vivo in the normal cellular context remains unclear. In the present study, we investigated the capacity of the sHsp, HspB8/Hsp22, to prevent protein aggregation in the cells using the polyglutamine protein Htt43Q as a model. In control conditions, Htt43Q accumulated in perinuclear inclusions composed of SDS-insoluble aggregates. Co-transfected with Htt43Q, HspB8 became occasionally trapped within the inclusions; however, in most cells, HspB8 blocked inclusion formation. Biochemical analyses indicated that HspB8 inhibited the accumulation of SDS-insoluble Htt43Q as efficiently as Hsp40 which was taken as a positive control. Htt43Q then accumulated in the SDS-soluble fraction, provided that protein degradation was blocked by proteasome and autophagy inhibitors. In contrast, the other sHsp Hsp27/HspB1 and alpha B-crystallin/HspB5 had no effect. This suggested that HspB8 functions as a molecular chaperone, maintaining Htt43Q in a soluble state competent for rapid degradation. Analyses of Hsp27-HspB8 chimeric proteins indicated that the C-terminal domain of HspB8 contains the specific sequence necessary for chaperone activity. Missense mutations in this domain at lysine 141, which are found in human motor neuropathies, significantly reduced the chaperone activity of the protein. A decrease in the HspB8 chaperone activity may therefore contribute to the development of these diseases.