Exogenous delivery of chaperonin subunit fragment ApiCCT1 modulates mutant Huntingtin cellular phenotypes
Exogenous delivery of chaperonin subunit fragment ApiCCT1 modulates mutant Huntingtin cellular phenotypes
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DOI：
10.1073/pnas.1222663110
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发表时间：
2013-02-19
影响因子：
11.1
通讯作者：
Thompson, Leslie M.
中科院分区：
文献类型：
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作者：
Sontag, Emily M.;Joachimiak, Lukasz A.;Thompson, Leslie M.
错误折叠蛋白的聚集是许多神经退行性疾病的特征，包括亨廷顿病 (HD)。 CCT/TRiC（含有 TCP-1/TCP-1 环的伴侣蛋白）伴侣蛋白复合物可以抑制由扩展的重复亨廷顿蛋白 (mHtt) 片段引起的聚集和细胞毒性。 CCT/TRiC 亚基 CCT1 的底物结合顶端结构域 ApiCCT1 足以在体外抑制扩展重复 mHtt 片段的聚集，为 HD 提供治疗前景。然而，考虑将 ApiCCT1 作为潜在治疗方法的一个关键障碍是交付。由于 ApiCCT1 具有与 HIV Tat 蛋白细胞转导结构域相似的区域，因此我们测试了重组 ApiCCT1 (ApiCCT1(r)) 蛋白是否可以在外源递送后进入细胞并调节一组已建立的 mHtt 介导的基于细胞的表型。细胞分级分离研究表明，外源 ApiCCT1(r) 可以穿透细胞膜并定位到细胞核，这与针对 HD 中细胞质和细胞核致病事件的策略一致。 ApiCCT1(r) 应用确实通过减少源自截短的 mHtt 外显子 1 片段表达的可见包涵体、纤维状寡聚体和不溶性 mHtt 的形成来调节 HD 细胞表型。通过实时成像观察，ApiCCT1(r) 还可以延迟包涵体形成的开始。 ApiCCT1(r) 降低了来自全长敲入 HD 小鼠的永生化纹状体细胞中 mHtt 介导的毒性，表明治疗益处可能超出了对聚集的影响。这些研究为针对 mHtt 介导的蛋白质发病机制的潜在稳健且独特的治疗策略提供了基础。
Aggregation of misfolded proteins is characteristic of a number of neurodegenerative diseases, including Huntington disease (HD). The CCT/TRiC (chaperonin containing TCP-1/TCP-1 ring) chaperonin complex can inhibit aggregation and cellular toxicity induced by expanded repeat Huntingtin (mHtt) fragments. The substrate-binding apical domain of CCT/TRiC subunit CCT1, ApiCCT1, is sufficient to inhibit aggregation of expanded repeat mHtt fragments in vitro, providing therapeutic promise for HD. However, a key hurdle in considering ApiCCT1 as a potential treatment is in delivery. Because ApiCCT1 has a region of similarity to the HIV Tat protein cell-transduction domain, we tested whether recombinant ApiCCT1 (ApiCCT1(r)) protein could enter cells following exogenous delivery and modulate an established panel of mHtt-mediated cell-based phenotypes. Cell fractionation studies demonstrate that exogenous ApiCCT1(r) can penetrate cell membranes and can localize to the nucleus, consistent with a strategy that can target both cytosolic and nuclear pathogenic events in HD. ApiCCT1(r) application does indeed modulate HD cellular phenotypes by decreasing formation of visible inclusions, fibrillar oligomers, and insoluble mHtt derived from expression of a truncated mHtt exon 1 fragment. ApiCCT1(r) also delays the onset of inclusion body formation as visualized via live imaging. ApiCCT1(r) reduces mHtt-mediated toxicity in immortalized striatal cells derived from full-length knock-in HD mice, suggesting that therapeutic benefit may extend beyond effects on aggregation. These studies provide the basis for a potentially robust and unique therapeutic strategy to target mHtt-mediated protein pathogenesis.