He Xiaoqing, Chen Michael G, Ma Qiang
Receptor Biology Laboratory, Toxicology and Molecular Biology Branch, Health Effects Laboratory Division, National Institute for Occupational Safety and Health, Centers for Disease Control and Prevention, Morgantown, West Virginia 26505, USA.
Chem Res Toxicol. 2008 Jul;21(7):1375-83. doi: 10.1021/tx800019a. Epub 2008 May 31.
Exposure to cadmium (Cd) elicits a range of adverse responses including oxidative damage and cancer. The molecular targets of Cd remain largely unidentified. Here, we analyzed the function and signal transduction of transcription factor Nrf2 in protection against Cd-induced oxidative stress. Wild-type (Nrf2 (+/+)) mouse embryonic fibroblasts (MEF) produced reactive oxygen species (ROS) at a low level, whereas treatment with Cd significantly increased the ROS production. On the other hand, Nrf2 knockout (Nrf2 (-/-)) MEF cells exhibited an elevated level of ROS under a basal condition, and Cd dramatically increased the ROS production at concentrations as low as 2 microM, resulting in increased sensitivity to Cd-induced cell death. Cd induced the basal and inducible expression of cytoprotective enzymes NQO1 and HO1 in WT MEF cells, but induction was lost in Nrf2 (-/-) MEF cells. Induction of the genes required antioxidant response elements (ARE) as Cd drove ARE-dependent reporter expression and Cd-activated Nrf2 bound to endogenous AREs in mouse hepa1c1c7 cells. Activation of Nrf2 by Cd involved stabilization of the Nrf2 protein, increased formation of Nrf2/Keap1 complex in the cytoplasm, translocation of the complex into the nucleus, and subsequently disruption of the complex. Lastly, Nrf2 was found ubiquitinated in the cytoplasm but deubiquitinated in the nucleus. The study provided a mechanistic transcriptional model in which Cd activates Nrf2 through a metal-activated signaling pathway involving a dynamic interplay between ubiquitination/deubiquitination and complex formation/dissociation of Nrf2 and Keap1.
接触镉（Cd）会引发一系列不良反应，包括氧化损伤和癌症。Cd的分子靶点在很大程度上仍未明确。在此，我们分析了转录因子Nrf2在抵御Cd诱导的氧化应激中的功能和信号转导。野生型（Nrf2(+/+)）小鼠胚胎成纤维细胞（MEF）产生的活性氧（ROS）水平较低，而用Cd处理会显著增加ROS的产生。另一方面，Nrf2基因敲除（Nrf2(-/-)）的MEF细胞在基础条件下ROS水平升高，并且Cd在低至2 microM的浓度下就能显著增加ROS的产生，导致对Cd诱导的细胞死亡敏感性增加。Cd诱导野生型MEF细胞中细胞保护酶NQO1和HO1的基础表达和诱导性表达，但在Nrf2(-/-)的MEF细胞中这种诱导作用消失。这些基因的诱导需要抗氧化反应元件（ARE），因为Cd驱动ARE依赖性报告基因表达，并且Cd激活的Nrf2与小鼠hepa1c1c7细胞中的内源性ARE结合。Cd对Nrf2的激活涉及Nrf2蛋白的稳定、细胞质中Nrf2/Keap1复合物形成增加、该复合物向细胞核的转位以及随后复合物的解离。最后，发现Nrf2在细胞质中被泛素化，但在细胞核中被去泛素化。该研究提供了一个机制转录模型，其中Cd通过涉及泛素化/去泛素化以及Nrf2和Keap1复合物形成/解离之间动态相互作用的金属激活信号通路激活Nrf2。
