Department of Biomedical Sciences, School of Veterinary Medicine, University of Pennsylvania, Philadelphia, PA, USA.
Department of Biophysics and Free Radical Research Center, Medical College of Wisconsin, Milwaukee, WI, USA.
Redox Biol. 2020 Sep;36:101606. doi: 10.1016/j.redox.2020.101606. Epub 2020 Jun 21.
The mitochondrial electron transport chain is a major source of reactive oxygen species (ROS) and is also a target of ROS, with an implied role in the stabilization of hypoxia-inducible factor (HIF) and induction of the AMPK pathway. Here we used varying doses of two agents, Mito-Paraquat and Mito-Metformin, that have been conjugated to cationic triphenylphosphonium (TPP) moiety to selectively target them to the mitochondrial matrix compartment, thereby resulting in the site-specific generation of ROS within mitochondria. These agents primarily induce superoxide (O) production by acting on complex I. In Raw264.7 macrophages, C2C12 skeletal myocytes, and HCT116 adenocarcinoma cells, we show that mitochondria-targeted oxidants can induce ROS (O and HO). In all three cell lines tested, the mitochondria-targeted agents disrupted membrane potential and activated calcineurin and the Cn-dependent retrograde signaling pathway. Hypoxic culture conditions also induced Cn activation and HIF1α activation in a temporally regulated manner, with the former appearing at shorter exposure times. Together, our results indicate that mitochondrial oxidant-induced retrograde signaling is driven by disruption of membrane potential and activation of Ca/Cn pathway and is independent of ROS-induced HIF1α or AMPK pathways.
线粒体电子传递链是活性氧（ROS）的主要来源，也是 ROS 的靶点，在稳定缺氧诱导因子（HIF）和诱导 AMPK 通路方面发挥作用。在这里，我们使用了两种不同剂量的试剂，即 Mito-paraquat 和 Mito-metformin，它们已与阳离子三苯基膦（TPP）部分缀合，以选择性地将它们靶向线粒体基质隔室，从而导致线粒体中 ROS 的位点特异性生成。这些试剂主要通过作用于复合物 I 来诱导超氧化物（O）的产生。在 Raw264.7 巨噬细胞、C2C12 骨骼肌细胞和 HCT116 腺癌细胞中，我们表明，靶向线粒体的氧化剂可以诱导 ROS（O 和 HO）。在所有三种测试的细胞系中，靶向线粒体的试剂破坏了膜电位并激活了钙调神经磷酸酶和 Cn 依赖性逆行信号通路。低氧培养条件也以时间调节的方式诱导 Cn 激活和 HIF1α 激活，前者出现在较短的暴露时间内。总之，我们的结果表明，线粒体氧化剂诱导的逆行信号是由膜电位破坏和 Ca/Cn 通路激活驱动的，与 ROS 诱导的 HIF1α 或 AMPK 通路无关。
