Effects of molecular chaperones on polyQ-mediated cell death and toxicity using cellular model of SBMA

1. 我们利用SBMA细胞模型研究了Hsp105α对多q束扩增引起的聚集体形成和细胞毒性的影响。含有扩大多q束的截断雄激素受体（tARs）的短暂表达诱导COS-7和SK-N-SH细胞聚集形成，并伴随这些核聚集细胞的凋亡细胞死亡。当Hsp105α与tAR97在细胞中过表达时，聚集体的形成和polyQ束扩张引起的细胞毒性明显降低。Hsp105α的β-片结构域和α-螺旋结构域是抑制体内外聚集体形成的必需结构域，而非atp酶结构域。此外，在SBMA患者和转基因小鼠的组织中，Hsp105α被发现定位于含有扩增多q束的Ars形成的核包裹体中。这些研究结果表明，Hsp105α的过表达可抑制无伴侣活性的多q束扩张引起的细胞死亡，并且在脑中增强Hsp105α必需结构域的表达可能为CAG重复疾病提供有效的治疗手段。我们研究了载体蛋白对聚q束扩张引起的聚集体形成和细胞毒性的影响。当在COS-7细胞中表达含有与GFP融合的扩增多q束（tAR24和tAR97）或与GFP融合的扩增多q束（polyQ24和polyQ97）的截断Ars时，polyQ97比tAR97引起明显的聚集形成和凋亡。此外，虽然tAR24和polyQ24都没有引起它们的聚集，但polyQ24而不是tAR24诱导细胞凋亡。因此，提示携带蛋白的多q束影响多q束的细胞毒性。蛋白质组学分析目前正在取得进展，以发现负责细胞毒性和细胞凋亡的蛋白质，这些蛋白质表达含有扩增多q束的tar。

1. We examined the effects of Hsp105α on the aggregate formation and cytotoxicity caused by expansion of polyQ tracts using cellular model of SBMA. The transient expression of truncated androgen receptors (tARs) containing expanded polyQ tracts induced aggregate formation in COS-7 and SK-N-SH cells and concomitantly apoptotic cell death in these cells with nuclear aggregates. When Hsp105α was over-expressed with tAR97 in cells, the aggregate formation and cytotoxicity caused by expansion of polyQ tracts were markedly reduced. Both β-sheet and α-helix domains, but not ATPase domain, of Hsp105α were necessary for the suppression of the aggregate formation in vivo and in vitro. Furthermore, Hsp105α was found to localize in nuclear inclusions formed by Ars containing expanded polyQ tracts in tissues of patients and transgenic mice with SBMA. These findings suggest that over-expression of Hsp105α suppresses cell death caused by expansion of polyQ tracts without chaperone activity, and the enhanced expression of the essential domains of Hsp105α in brain may provide an effective therapeutic mean for CAG repeat diseases.2. We examined the effect of carrier protein on the aggregate formation and cytotoxicity caused by expansion of polyQ tracts. When truncated Ars containing expanded polyQ tracts fused to GFP (tAR24 and tAR97) or expanded polyQ tracts fused to GFP (polyQ24 and polyQ97) were expressed in COS-7 cells, polyQ97 caused marked aggregate formation and apoptosis than tAR97 did. Furthermore, although both tAR24 and polyQ24 did not cause their aggregation, polyQ24 but not tAR24 induced apoptosis. Thus, it was suggested that protein carrying polyQ tracts affects the cytotoxicity of polyQ tract.3. Proteome analysis is now progress to find proteins which are responsible for cytotoxicity and apoptosis of cells expressing tARs containing expanded polyQ tracts.