Abboud Cardiovascular Research Center, Division of Cardiovascular Medicine, Department of Internal Medicine, Carver College of Medicine, University of Iowa, Iowa City, IA 52242, USA.
Free Radical and Radiation Biology Program, Department of Radiation Oncology, Holden Comprehensive Cancer Center, University of Iowa, Iowa City, IA 52242, USA.
Redox Biol. 2018 Jun;16:401-413. doi: 10.1016/j.redox.2018.04.001. Epub 2018 Apr 3.
Reduction of oxidized methionines is emerging as a major protein repair pathway. The lack of methionine sulfoxide reductase A (MsrA) exacerbates cardiovascular disease phenotypes driven by increased oxidative stress. However, the role of MsrA on maintaining cellular homeostasis in the absence of excessive oxidative stress is less well understood.
Constitutive genetic deletion of MsrA increased formation of p62-containing protein aggregates, activated autophagy, and decreased a marker of apoptosis in vascular smooth muscle cells (VSMC). The association of Keap1 with p62 was augmented in MsrA-/- VSMC. Keap1 targets the transcription factor Nrf2, which regulates antioxidant genes, for proteasomal degradation. However, in MsrA-/- VSMC, the association of Nrf2 with Keap1 was diminished. Whereas Nrf2 mRNA levels were not decreased in MsrA-/- VSMC, we detected decreased ubiquitination of Nrf2 and a corresponding increase in total Nrf2 protein in the absence of biochemical markers of oxidative stress. Moreover, nuclear-localized Nrf2 was increased under MsrA deficiency, resulting in upregulation of Nrf2-dependent transcriptional activity. Consequently, transcription, protein levels and enzymatic activity of glutamate-cysteine ligase and glutathione reductase were greatly augmented in MsrA-/- VSMC.
Our findings demonstrate that reversal of methionine oxidation is required for maintenance of cellular homeostasis in the absence of increased oxidative stress. These data provide the first link between autophagy and activation of Nrf2 in the setting of MsrA deletion.
氧化甲硫氨酸的还原作用正在成为一种主要的蛋白质修复途径。缺乏甲硫氨酸亚砜还原酶 A（MsrA）会加剧由氧化应激增加引起的心血管疾病表型。然而，在没有过度氧化应激的情况下，MsrA 对维持细胞内稳态的作用还不太清楚。
MsrA 的组成型遗传缺失会增加含 p62 的蛋白质聚集体的形成，激活自噬，并降低血管平滑肌细胞（VSMC）中凋亡的标志物。在 MsrA-/-VSMC 中，Keap1 与 p62 的结合增强。Keap1 将转录因子 Nrf2 靶向蛋白酶体降解，Nrf2 调节抗氧化基因。然而，在 MsrA-/-VSMC 中，Nrf2 与 Keap1 的结合减少。尽管 MsrA-/-VSMC 中的 Nrf2 mRNA 水平没有降低，但我们检测到 Nrf2 的泛素化减少，并且在没有生化氧化应激标志物的情况下，总 Nrf2 蛋白相应增加。此外，在 MsrA 缺乏的情况下，核定位的 Nrf2 增加，导致 Nrf2 依赖性转录活性上调。因此，谷氨酸半胱氨酸连接酶和谷胱甘肽还原酶的转录、蛋白水平和酶活性在 MsrA-/-VSMC 中大大增加。
我们的研究结果表明，在没有增加氧化应激的情况下，甲硫氨酸氧化的逆转对于维持细胞内稳态是必需的。这些数据提供了 MsrA 缺失时自噬与 Nrf2 激活之间的第一个联系。
