Cytoplasmic Inclusions of Htt Exon1 Containing an Expanded Polyglutamine Tract Suppress Execution of Apoptosis in Sympathetic Neurons
Cytoplasmic Inclusions of Htt Exon1 Containing an Expanded Polyglutamine Tract Suppress Execution of Apoptosis in Sympathetic Neurons
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DOI：
10.1523/jneurosci.4751-08.2008
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发表时间：
2008-12-31
影响因子：
5.3
通讯作者：
Tolkovsky, Aviva M.
中科院分区：
文献类型：
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作者：
King, Matthew A.;Goemans, Christoph G.;Tolkovsky, Aviva M.
含有延长的多聚谷氨酰胺重复序列的蛋白质导致至少九种神经退行性疾病，但疾病相关的神经元死亡的机制仍然不确定。我们发现，交感神经元含有由97个谷氨酰胺内表达的人类亨廷顿蛋白外显子1增强的绿色荧光蛋白（Q97）形成的细胞质内含物进行了长期形式的非凋亡性死亡是不敏感的Bax缺失或caspase抑制，但其特征是线粒体功能障碍。通过联合阿糖胞苷和神经生长因子撤退治疗神经元，我们证明，Q97赋予强大的抗凋亡在多个水平：尽管正常的促凋亡信号（升高P-ser 15-p53和BimEL），有没有增加的彪马mRNA或Bax激活，都是必要的凋亡。即使用过表达的Puma恢复Bax易位也不会激活凋亡。我们证明，这种强大的抑制细胞凋亡是由Q97介导的积累的热休克蛋白70，这是通过抑制蛋白酶体活性。因此，细胞凋亡通过短发夹RNA介导的Hsp 70敲低而恢复。这些发现解释了在表达Q97的神经元中凋亡性死亡的罕见性。考虑到蛋白酶体阻断，我们测试用雷帕霉素增强溶酶体介导的降解是否减少Q97积累。雷帕霉素使非病理性Q25的量在3天内减少70%，但Q97的积累不受影响。有趣的是，由于组成性溶酶体降解，Q47包涵体形成得更慢，但更快形成的Q97包涵体逃脱了溶酶体的控制。因此，细胞质Q97夹杂物是难清除的蛋白酶体和溶酶体系统，导致的毒性，占主导地位的神经保护热休克蛋白70。我们的研究结果可以解释细胞凋亡的罕见性，但与polyQ包涵体疾病相关的不可避免的细胞死亡。
Proteins containing extended polyglutamine repeats cause at least nine neurodegenerative disorders, but the mechanisms of disease-related neuronal death remain uncertain. We show that sympathetic neurons containing cytoplasmic inclusions formed by 97 glutamines expressed within human huntingtin exon1 - enhanced green fluorescent protein (Q97) undergo a protracted form of nonapoptotic death that is insensitive to Bax deletion or caspase inhibition but is characterized by mitochondrial dysfunction. By treating the neurons with combined cytosine arabinoside and NGF withdrawal, we demonstrate that Q97 confers a powerful resistance to apoptosis at multiple levels: despite normal proapoptotic signaling ( elevation of P-ser15-p53 and BimEL), there is no increase of Puma mRNA or Bax activation, both necessary for apoptosis. Even restoration of Bax translocation with overexpressed Puma does not activate apoptosis. We demonstrate that this robust inhibition of apoptosis is caused by Q97-mediated accumulation of Hsp70, which occurs through inhibition of proteasomal activity. Thus, apoptosis is reinstated by short hairpin RNA-mediated knockdown of Hsp70. These findings explain the rarity of apoptotic death in Q97-expressing neurons. Given the proteasomal blockade, we test whether enhancing lysosomal-mediated degradation with rapamycin reduces Q97 accumulation. Rapamycin reduces the amount of nonpathological Q25 by 70% over 3 d, but Q97 accumulation is unaffected. Interestingly, Q47 inclusions form more slowly as a result of constitutive lysosomal degradation, but faster-forming Q97 inclusions escape lysosomal control. Thus, cytoplasmic Q97 inclusions are refractory to clearance by proteasomal and lysosomal systems, leading to a toxicity that dominates over neuroprotective Hsp70. Our findings may explain the rarity of apoptosis but the inevitable cell death associated with polyQ inclusion diseases.