Wichmann Anita E, Thomson Nicole M, Peterson Lisa A, Wattenberg Elizabeth V
Division of Environmental and Occupational Health, School of Public Health, University of Minnesota, Minneapolis, Minnesota 55455, USA.
Chem Res Toxicol. 2003 Jan;16(1):87-94. doi: 10.1021/tx0256026.
Mitogen-activated protein kinases (MAPKs) play a central role in transmitting stress-induced signals stimulated by genotoxic agents. The present study is the first to investigate the mechanisms by which genotoxic alkylating agents modulate MAPKs by directly measuring the effects of methylating agents on MAPK activity, DNA methylation, and intracellular glutathione levels. The effects of acetoxymethylmethylnitrosamine (AMMN), N-nitroso-N-methylurethane (NMUR), and N-methyl-N-nitrosourea (MNU) on these parameters were compared in a fetal rat lung cell line model (MP48). These compounds were chosen because they methylate DNA via a methanediazonium intermediate and, therefore, should induce similar cellular methylation patterns, although they produce different side products upon decomposition. All three compounds stimulated the activation of the stress-activated MAPKs, c-Jun N-terminal kinase, and p38. In contrast to what has been reported for other methylating agents, these compounds also stimulated the activation of extracellular signal regulated kinase (ERK), a MAPK typically activated by mitogenic agents. O6-methylguanine (O6-mG) is widely considered to be the critical toxic lesion induced by methylating agents, including AMMN, NMUR, and MNU, which form DNA adducts through SN1 reactions. O6-mG does not appear to be a key regulator of MAPK activity by these compounds, however. There is no direct relationship between the levels of O6-mG and the levels of MAPK activation, and formation of O6-mG does not appear to be sufficient to stimulate MAPK activation. The present studies also indicate that depletion of glutathione is not required or sufficient to stimulate MAPK activation by the methylating agents investigated here. The use of a pharmacological inhibitor indicates that these methylating agents activate ERK through a signaling pathway that requires the ERK kinase MEK. Altogether, these data indicate that genotoxic methylating agents activate MAPKs through mechanisms that are likely to involve the alkylation of cellular targets other than DNA.
丝裂原活化蛋白激酶（MAPKs）在传递遗传毒性剂刺激的应激诱导信号中起核心作用。本研究首次通过直接测量甲基化剂对MAPK活性、DNA甲基化和细胞内谷胱甘肽水平的影响，来探究遗传毒性烷基化剂调节MAPKs的机制。在胎鼠肺细胞系模型（MP48）中比较了乙酰氧甲基甲基亚硝胺（AMMN）、N-亚硝基-N-甲基脲（NMUR）和N-甲基-N-亚硝基脲（MNU）对这些参数的影响。选择这些化合物是因为它们通过甲二氮鎓中间体使DNA甲基化，因此，尽管它们在分解时会产生不同的副产物，但应该会诱导相似的细胞甲基化模式。所有这三种化合物均刺激了应激激活的MAPKs、c-Jun氨基末端激酶和p38的活化。与其他甲基化剂的报道不同，这些化合物还刺激了细胞外信号调节激酶（ERK）的活化，ERK是一种通常由促有丝分裂剂激活的MAPK。O6-甲基鸟嘌呤（O6-mG）被广泛认为是由甲基化剂诱导的关键毒性损伤，包括AMMN、NMUR和MNU，它们通过SN1反应形成DNA加合物。然而，O6-mG似乎不是这些化合物调节MAPK活性的关键调节因子。O6-mG的水平与MAPK活化水平之间没有直接关系，并且O6-mG的形成似乎不足以刺激MAPK活化。本研究还表明，谷胱甘肽的消耗对于此处研究甲基化剂刺激MAPK活化既不是必需的也不是充分的。使用药理学抑制剂表明，这些甲基化剂通过需要ERK激酶MEK的信号通路激活ERK。总之，这些数据表明，遗传毒性甲基化剂通过可能涉及DNA以外的细胞靶点烷基化的机制激活MAPKs。
