Department of Pharmacology & Toxicology, College of Pharmacy, University of Arizona, Tucson, AZ, USA.
PLoS One. 2009 Aug 11;4(8):e6588. doi: 10.1371/journal.pone.0006588.
The bZIP transcription factor Nrf2 has emerged as a pivotal regulator of intracellular redox homeostasis by controlling the expression of many endogenous antioxidants and phase II detoxification enzymes. Upon oxidative stress, Nrf2 is induced at protein levels through redox-sensitive modifications on cysteine residues of Keap1, a component of the E3 ubiquitin ligase that targets Nrf2 for ubiquitin-dependent degradation. The mitogen activated protein kinases (MAPKs) have previously been proposed to regulate Nrf2 in response to oxidative stress. However, the exact role of MAPKs and the underlying molecular mechanism remain poorly defined. Here we report the first evidence that Nrf2 is phosphorylated in vivo by MAPKs. We have identified multiple serine/threonine residues as major targets of MAPK-mediated phosphorylation. Combined alanine substitution on those residues leads to a moderate decrease in the transcriptional activity of Nrf2, most likely due to a slight reduction in its nuclear accumulation. More importantly, Nrf2 protein stability, primarily controlled by Keap1, is not altered by Nrf2 phosphorylation in vivo. These data indicate that direct phosphorylation of Nrf2 by MAPKs has limited contribution in modulating Nrf2 activity. We suggest that MAPKs regulate the Nrf2 signaling pathway mainly through indirect mechanisms.
bZIP 转录因子 Nrf2 通过控制许多内源性抗氧化剂和 II 相解毒酶的表达，成为细胞内氧化还原稳态的关键调节剂。在氧化应激下，Nrf2 通过 Keap1 上半胱氨酸残基的氧化还原敏感修饰在蛋白质水平上被诱导，Keap1 是一种 E3 泛素连接酶的组成部分，该酶将 Nrf2 作为泛素依赖性降解的靶标。丝裂原活化蛋白激酶（MAPKs）先前被提议在氧化应激时调节 Nrf2。然而，MAPKs 的确切作用及其潜在的分子机制仍未得到充分定义。在这里，我们首次报道了 MAPK 体内磷酸化 Nrf2 的证据。我们已经确定了多个丝氨酸/苏氨酸残基作为 MAPK 介导的磷酸化的主要靶标。这些残基的组合丙氨酸取代导致 Nrf2 的转录活性适度降低，这很可能是由于其核积累略有减少。更重要的是，Nrf2 蛋白稳定性主要由 Keap1 控制，在体内 Nrf2 磷酸化并不改变。这些数据表明，MAPK 对 Nrf2 的直接磷酸化在调节 Nrf2 活性方面的贡献有限。我们认为，MAPKs 主要通过间接机制调节 Nrf2 信号通路。
