Department of Cardiology, Keio University School of Medicine, Tokyo, Japan.
Department of Cardiology, Keio University School of Medicine, Tokyo, Japan.
J Mol Cell Cardiol. 2021 Dec;161:116-129. doi: 10.1016/j.yjmcc.2021.08.006. Epub 2021 Aug 12.
MITOL/MARCH5 is an E3 ubiquitin ligase that plays a crucial role in the control of mitochondrial quality and function. However, the significance of MITOL in cardiomyocytes under physiological and pathological conditions remains unclear. First, to determine the significance of MITOL in unstressed hearts, we assessed the cellular changes with the reduction of MITOL expression by siRNA in neonatal rat primary ventricular cardiomyocytes (NRVMs). MITOL knockdown in NRVMs induced cell death via ferroptosis, a newly defined non-apoptotic programmed cell death, even under no stress conditions. This phenomenon was observed only in NRVMs, not in other cell types. MITOL knockdown markedly reduced mitochondria-localized GPX4, a key enzyme associated with ferroptosis, promoting accumulation of lipid peroxides in mitochondria. In contrast, the activation of GPX4 in MITOL knockdown cells suppressed lipid peroxidation and cell death. MITOL knockdown reduced the glutathione/oxidized glutathione (GSH/GSSG) ratio that regulated GPX4 expression. Indeed, the administration of GSH or N-acetylcysteine improved the expression of GPX4 and viability in MITOL-knockdown NRVMs. MITOL-knockdown increased the expression of the glutathione-degrading enzyme, ChaC glutathione-specific γ-glutamylcyclotransferase 1 (Chac1). The knockdown of Chac1 restored the GSH/GSSG ratio, GPX4 expression, and viability in MITOL-knockdown NRVMs. Further, in cultured cardiomyocytes stressed with DOX, both MITOL and GPX4 were reduced, whereas forced-expression of MITOL suppressed DOX-induced ferroptosis by maintaining GPX4 content. Additionally, MITOL knockdown worsened vulnerability to DOX, which was almost completely rescued by treatment with ferrostatin-1, a ferroptosis inhibitor. In vivo, cardiac-specific depletion of MITOL did not produce obvious abnormality, but enhanced susceptibility to DOX toxicity. Finally, administration of ferrostatin-1 suppressed exacerbation of DOX-induced myocardial damage in MITOL-knockout hearts. The present study demonstrates that MITOL determines the cell fate of cardiomyocytes via the ferroptosis process and plays a key role in regulating vulnerability to DOX treatment. (288/300).
MITOL/MARCH5 是一种 E3 泛素连接酶，在控制线粒体质量和功能方面发挥着关键作用。然而，MITOL 在生理和病理条件下的心肌细胞中的意义尚不清楚。首先，为了确定 MITOL 在未受应激的心脏中的意义，我们通过 siRNA 降低了新生大鼠原代心室心肌细胞 (NRVM) 中的 MITOL 表达，评估了细胞变化。MITOL 敲低在没有应激条件下诱导心肌细胞通过铁死亡（一种新定义的非凋亡程序性细胞死亡）发生细胞死亡。这种现象仅在 NRVM 中观察到，而在其他细胞类型中未观察到。MITOL 敲低显著降低了定位于线粒体的 GPX4，这是一种与铁死亡相关的关键酶，促进了脂质过氧化物在线粒体中的积累。相比之下，在 MITOL 敲低细胞中激活 GPX4 抑制了脂质过氧化和细胞死亡。MITOL 敲低降低了调节 GPX4 表达的谷胱甘肽/氧化型谷胱甘肽 (GSH/GSSG) 比。事实上，给予 GSH 或 N-乙酰半胱氨酸可改善 MITOL 敲低 NRVM 中的 GPX4 表达和活力。MITOL 敲低增加了谷胱甘肽降解酶 ChaC 谷胱甘肽特异性γ-谷氨酰环转移酶 1 (Chac1) 的表达。Chac1 的敲低恢复了 MITOL 敲低 NRVM 中的 GSH/GSSG 比、GPX4 表达和活力。此外，在 DOX 应激的培养心肌细胞中，MITOL 和 GPX4 均减少，而强制表达 MITOL 通过维持 GPX4 含量抑制 DOX 诱导的铁死亡。此外，MITOL 敲低使 DOX 易感性恶化，而用铁死亡抑制剂 ferrostatin-1 处理几乎完全挽救了这种情况。在体内，心脏特异性敲低 MITOL 不会产生明显的异常，但增强了对 DOX 毒性的易感性。最后，给予 ferrostatin-1 抑制了 MITOL 敲除心脏中 DOX 诱导的心肌损伤的恶化。本研究表明，MITOL 通过铁死亡过程决定心肌细胞的命运，并在调节 DOX 治疗的易感性方面发挥关键作用。(288/300)。
