APOL1 Kidney Risk Variants Induce Cell Death via Mitochondrial Translocation and Opening of the Mitochondrial Permeability Transition Pore.
APOL1 Kidney Risk Variants Induce Cell Death via Mitochondrial Translocation and Opening of the Mitochondrial Permeability Transition Pore.
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DOI：
10.1681/asn.2019020114
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发表时间：
2019-09
期刊：
影响因子：
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通讯作者：
Shrijal S Shah;H. Lannon;Leny Dias;Jia-yue Zhang;S. Alper;M. Pollak;D. Friedman
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文献类型：
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作者：
Shrijal S Shah;H. Lannon;Leny Dias;Jia-yue Zhang;S. Alper;M. Pollak;D. Friedman
载脂蛋白L1（APOL 1）的遗传变异与非裔美国人CKD发病率的大幅增加有关。在细胞和小鼠模型中的实验表明，这些风险相关的多态性是毒性的功能获得性变体，在隐性遗传模式下导致肾功能障碍。最近在锥虫和人类细胞中的数据表明，这些变体可能通过其对线粒体的影响而引起毒性。方法为了研究APOL 1风险变异体诱导线粒体功能障碍的分子机制，我们产生了稳定表达APOL 1非风险G 0变异体或APOL 1风险变异体的四环素诱导的HEK 293 T-REx细胞。使用这些细胞，我们映射的分子途径，从线粒体进口的APOL 1蛋白的APOL 1诱导的细胞死亡与小干扰RNA敲低，药理学抑制剂，蓝色的天然PAGE，质谱，和线粒体通透性转换孔功能的评估。结果发现APOL 1 G 0和Risk变异蛋白进入线粒体基质的途径相同。一旦进入，G 0保持单体，而风险变体蛋白质倾向于形成更高阶的寡聚体。非风险G 0和风险变异蛋白结合的线粒体渗透性转换孔的组件，但只有风险变异蛋白激活孔开放。阻断APOL 1风险变体的线粒体输入在很大程度上消除了寡聚体形成，也挽救了毒性。结论：我们的研究阐明了APOL 1非风险和风险变体的分子行为的重要差异，我们的观察结果表明，尽管缺乏一致观察到的运输模式，细胞内定位或结合伴侣的差异，但可能解释这些变体非常不同的功能效应的机制。寡聚化模式的变异依赖性差异可能是APOL 1的隐性、功能获得性生物学的基础。
BACKGROUND Genetic Variants in Apolipoprotein L1 (APOL1) are associated with large increases in CKD rates among African Americans. Experiments in cell and mouse models suggest that these risk-related polymorphisms are toxic gain-of-function variants that cause kidney dysfunction, following a recessive mode of inheritance. Recent data in trypanosomes and in human cells indicate that such variants may cause toxicity through their effects on mitochondria. METHODS To examine the molecular mechanisms underlying APOL1 risk variant-induced mitochondrial dysfunction, we generated tetracycline-inducible HEK293 T-REx cells stably expressing the APOL1 nonrisk G0 variant or APOL1 risk variants. Using these cells, we mapped the molecular pathway from mitochondrial import of APOL1 protein to APOL1-induced cell death with small interfering RNA knockdowns, pharmacologic inhibitors, blue native PAGE, mass spectrometry, and assessment of mitochondrial permeability transition pore function. RESULTS We found that the APOL1 G0 and risk variant proteins shared the same import pathway into the mitochondrial matrix. Once inside, G0 remained monomeric, whereas risk variant proteins were prone to forming higher-order oligomers. Both nonrisk G0 and risk variant proteins bound components of the mitochondrial permeability transition pore, but only risk variant proteins activated pore opening. Blocking mitochondrial import of APOL1 risk variants largely eliminated oligomer formation and also rescued toxicity. CONCLUSIONS Our study illuminates important differences in the molecular behavior of APOL1 nonrisk and risk variants, and our observations suggest a mechanism that may explain the very different functional effects of these variants, despite the lack of consistently observed differences in trafficking patterns, intracellular localization, or binding partners. Variant-dependent differences in oligomerization pattern may underlie APOL1's recessive, gain-of-function biology.