Glyceraldehyde-3-phosphate Dehydrogenase (GAPDH) Aggregation Causes Mitochondrial Dysfunction during Oxidative Stress-induced Cell Death.
Glyceraldehyde-3-phosphate Dehydrogenase (GAPDH) Aggregation Causes Mitochondrial Dysfunction during Oxidative Stress-induced Cell Death.
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DOI：
10.1074/jbc.m116.759084
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发表时间：
2017-03-17
期刊：
影响因子：
--
通讯作者：
Takeuchi T
中科院分区：
文献类型：
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作者：
Nakajima H;Itakura M;Kubo T;Kaneshige A;Harada N;Izawa T;Azuma YT;Kuwamura M;Yamaji R;Takeuchi T
糖酵解甘油醛-3-磷酸脱氢酶（GAPDH）是一种多功能蛋白，也介导氧化应激下的细胞死亡。我们以前报道，GAPDH的活性位点半胱氨酸（Cys-152）在氧化应激诱导的GAPDH聚集与细胞死亡中起着至关重要的作用，C152 A-GAPDH突变体通过干扰野生型（WT）-GAPDH的聚集来挽救一氧化氮（NO）诱导的细胞死亡。然而，GAPDH聚集体诱导细胞死亡的详细机制仍然难以捉摸。在这里，我们报告说，NO诱导的GAPDH聚集专门导致线粒体功能障碍。首先，我们观察到NO诱导的GAPDH聚集和线粒体功能障碍之间的相关性，当GAPDH聚集发生在SH-SY 5 Y细胞的线粒体。在分离的线粒体中，WT-GAPDH的聚集体直接诱导线粒体肿胀和去极化，而含有C152 A-GAPDH的聚集体的混合物减少线粒体功能障碍。此外，用环孢菌素A治疗改善了WT-GAPDH聚集体诱导的肿胀和去极化。在多西环素诱导的SH-SY 5 Y细胞中，WT-GAPDH的过表达增强了NO诱导的线粒体功能障碍并增加了线粒体GAPDH聚集，而诱导的C152 A-GAPDH的过表达显著抑制了线粒体损伤。此外，NO诱导的细胞色素c释放到胞质溶胶和核转位的凋亡诱导因子从线粒体都增加在细胞过表达WT-GAPDH，但改善C152 A-GAPDH过表达细胞。有趣的是，GAPDH聚集体通过渗透性转换孔（PTP）开放诱导坏死细胞死亡。WT-或C152 A-GAPDH的表达不影响与蛋白质聚集相关的其他细胞死亡途径，如蛋白酶体抑制、内质网应激诱导的基因表达或自噬。总的来说，这些结果表明NO诱导的GAPDH聚集通过PTP开放特异性诱导线粒体功能障碍，导致细胞死亡。
Glycolytic glyceraldehyde-3-phosphate dehydrogenase (GAPDH) is a multifunctional protein that also mediates cell death under oxidative stress. We reported previously that the active-site cysteine (Cys-152) of GAPDH plays an essential role in oxidative stress-induced aggregation of GAPDH associated with cell death, and a C152A-GAPDH mutant rescues nitric oxide (NO)-induced cell death by interfering with the aggregation of wild type (WT)-GAPDH. However, the detailed mechanism underlying GAPDH aggregate-induced cell death remains elusive. Here we report that NO-induced GAPDH aggregation specifically causes mitochondrial dysfunction. First, we observed a correlation between NO-induced GAPDH aggregation and mitochondrial dysfunction, when GAPDH aggregation occurred at mitochondria in SH-SY5Y cells. In isolated mitochondria, aggregates of WT-GAPDH directly induced mitochondrial swelling and depolarization, whereas mixtures containing aggregates of C152A-GAPDH reduced mitochondrial dysfunction. Additionally, treatment with cyclosporin A improved WT-GAPDH aggregate-induced swelling and depolarization. In doxycycline-inducible SH-SY5Y cells, overexpression of WT-GAPDH augmented NO-induced mitochondrial dysfunction and increased mitochondrial GAPDH aggregation, whereas induced overexpression of C152A-GAPDH significantly suppressed mitochondrial impairment. Further, NO-induced cytochrome c release into the cytosol and nuclear translocation of apoptosis-inducing factor from mitochondria were both augmented in cells overexpressing WT-GAPDH but ameliorated in C152A-GAPDH-overexpressing cells. Interestingly, GAPDH aggregates induced necrotic cell death via a permeability transition pore (PTP) opening. The expression of either WT- or C152A-GAPDH did not affect other cell death pathways associated with protein aggregation, such as proteasome inhibition, gene expression induced by endoplasmic reticulum stress, or autophagy. Collectively, these results suggest that NO-induced GAPDH aggregation specifically induces mitochondrial dysfunction via PTP opening, leading to cell death.