Guyton K Z, Liu Y, Gorospe M, Xu Q, Holbrook N J
Section on Gene Expression and Aging, Gerontology Research Center, NIA, National Institutes of Health, Baltimore, Maryland 21224, USA.
J Biol Chem. 1996 Feb 23;271(8):4138-42. doi: 10.1074/jbc.271.8.4138.
The mitogen-activated protein kinase (MAPK) family is comprised of key regulatory proteins that control the cellular response to both proliferation and stress signals. In this study we investigated the factors controlling MAPK activation by H2O2 and explored the impact of altering the pathways to kinase activation on cell survival following H2O2 exposure. Potent activation (10-20-fold) of extracellular signal-regulated protein kinase (ERK2) occurred within 10 min of H2O2 treatment, whereupon rapid inactivation ensued. H2O2 activated ERK2 in several cell types and also moderately activated (3-5-fold) both c-Jun N-terminal kinase and p38/RK/CSBP. Additionally, H2O2 increased the mRNA expression of MAPK-dependent genes c-jun, c-fos, and MAPK phosphatase-1. Suramin pretreatment completely inhibited H2O2 stimulation of ERK2, highlighting a role for growth factor receptors in this activation. Further, ERK2 activation by H2O2 was blocked by pretreatment with either N-acetyl-cysteine, o-phenanthroline, or mannitol, indicating that metal-catalyzed free radical formation mediates the initiation of signal transduction by H2O2. H2O2-stimulated activation of ERK2 was abolished in PC12 cells by inducible or constitutive expression of the dominant negative Ras-N-17 allele. Interestingly, PC12/Ras-N-17 cells were more sensitive than wild-type PC12 cells to H2O2 toxicity. Moreover, NIH 3T3 cells expressing constitutively active MAPK kinase (MEK, the immediate upstream regulator of ERK) were more resistant to H2O2 toxicity, while those expressing kinase-defective MEK were more sensitive, than cells expressing wild-type MEK. Taken together, these studies provide insight into mechanisms of MAPK regulation by H2O2 and suggest that ERK plays a critical role in cell survival following oxidant injury.
丝裂原活化蛋白激酶（MAPK）家族由关键调节蛋白组成，这些蛋白控制细胞对增殖和应激信号的反应。在本研究中，我们调查了控制H2O2激活MAPK的因素，并探讨了改变激酶激活途径对H2O2暴露后细胞存活的影响。H2O2处理后10分钟内，细胞外信号调节蛋白激酶（ERK2）发生强力激活（10 - 20倍），随后迅速失活。H2O2在几种细胞类型中激活ERK2，同时也适度激活（3 - 5倍）c-Jun N末端激酶和p38/RK/CSBP。此外，H2O2增加了MAPK依赖性基因c-jun、c-fos和MAPK磷酸酶-1的mRNA表达。苏拉明预处理完全抑制了H2O2对ERK2的刺激，突出了生长因子受体在这种激活中的作用。此外，用N-乙酰半胱氨酸、邻菲罗啉或甘露醇预处理可阻断H2O2对ERK2的激活，表明金属催化的自由基形成介导了H2O2信号转导的起始。通过诱导或组成型表达显性负性Ras-N-17等位基因，H2O2刺激的ERK2激活在PC12细胞中被消除。有趣的是，PC12/Ras-N-17细胞比野生型PC12细胞对H2O2毒性更敏感。此外，组成型表达活性MAPK激酶（MEK，ERK的直接上游调节因子）的NIH 3T3细胞比表达野生型MEK的细胞对H2O2毒性更具抗性，而表达激酶缺陷型MEK的细胞则更敏感。综上所述，这些研究深入了解了H2O2对MAPK的调节机制，并表明ERK在氧化损伤后的细胞存活中起关键作用。
