Comparison of p53 mutations induced by PAH o-quinones with those caused by anti-benzo[a] pyrene diol epoxide in vitro:: Role of reactive oxygen and biological selection
Comparison of p53 mutations induced by PAH o-quinones with those caused by anti-benzo[a] pyrene diol epoxide in vitro:: Role of reactive oxygen and biological selection
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DOI：
10.1021/tx0601206
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发表时间：
2006-11-20
影响因子：
4.1
通讯作者：
Field, Jeffrey
中科院分区：
文献类型：
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作者：
Shen, Yu-Min;Troxel, Andrea B.;Field, Jeffrey
多环芳烃（PAH）是烟草烟雾中的主要致癌物质之一。它们通过不同的途径代谢激活，形成二醇环氧化物、多环芳烃邻醌或自由基阳离子，其中每一种都被认为是最终致癌物。为了研究 PAH 代谢物如何突变 p53，我们使用了基于 p53 转录活性的酵母报告基因测定。表达野生型 p53 的集落变白 (ADE+)，表达突变型 p53 的集落变红 (ADE-)。我们检查了三种邻醌苯并[a]芘-7,8-二酮、苯并[a]蒽-3,4-二酮和二甲基苯并[a]蒽-3,4-二酮的致突变性，并将它们与同一系统内的(+/-)-抗苯并[a]芘二醇环氧化物((+/-)-抗BPDE)进行了比较。如果使用氧化还原循环条件，测试的 PAH 邻醌突变频率呈剂量依赖性增加，范围为 0.160-0.375 μM 醌。显性突变是 G 到 T 颠换（> 42%），DNA 结合域中热点突变的发生率是随机分布预期的两倍多。 G 到 T 颠换对氧化还原循环的依赖性表明 8-oxo-dGuo 是造成损伤的原因，它是在相同条件下产生的（Chem. Res. Toxicol. (2005) 18, 1027）。使用(+/-)-抗-BPDE也观察到剂量依赖性突变频率，但在微摩尔浓度(0-20μM)下。观察到的突变模式在未甲基化 p53 中为 G 到 C (63%) > G 到 A (18%) > G 到 T (15%)，在甲基化 p53 中为 G 到 A (39%) > G 到 C (34%) > G 到 T (16%)。 G 突变的优势与抗 BPDE-N-2-dGuo 作为主要加合物的形成一致。 (+/-)-抗-BPDE 突变的热点频率在未甲基化和甲基化 p53 中基本上是随机的，表明 5'-CpG-3' 岛在测定中并不直接突变。这些数据表明，吸烟可能会形成 PAH 邻醌，从而产生活性氧，从而导致 p53 突变。由此产生的 8-oxo-dGuo 产生了一种突变模式，但与肺癌中观察到的突变谱不一致；我们认为光谱的出现需要生物选择。
Polycyclic aromatic hydrocarbons (PAH) are one of the major carcinogens in tobacco smoke. They are metabolically activated through different routes to form either diol-epoxides, PAH o-quinones, or radical cations, each of which has been proposed to be an ultimate carcinogen. To study how PAH metabolites mutate p53, we used a yeast reporter gene assay based on p53 transcriptional activity. Colonies expressing wt p53 turn white (ADE+) and those expressing mutant p53 turn red (ADE-). We examined the mutagenicity of three o-quinones, benzo[a]pyrene-7,8-dione, benz[a]anthracene-3,4-dione, and dimethylbenz[a]anthracene-3,4-dione, and compared them with (+/-)-anti-benzo[a]pyrene diol epoxide ((+/-)-anti-BPDE) within the same system. The PAH o-quinones tested gave a dose-dependent increase in mutation frequency in the range of 0.160-0.375 mu M quinone, provided redox-cycling conditions were used. The dominant mutations were G to T transversions (> 42%), and the incidence of hotspot mutations in the DNA-binding domain was more than twice than that expected by a random distribution. The dependence of G to T transversions on redox cycling implicates 8-oxo-dGuo as the lesion responsible, which is produced under identical conditions (Chem. Res. Toxicol. (2005) 18, 1027). A dose-dependent mutation frequency was also observed with (+/-)-anti-BPDE but at micromolar concentrations (0-20 mu M). The mutation pattern observed was G to C (63%) > G to A (18%) > G to T (15%) in umethylated p53 and was G to A (39%) > G to C (34%) > G to T (16%) in methylated p53. The preponderance of G mutations is consistent with the formation of anti-BPDE-N-2-dGuo as the major adduct. The frequency of hotspots mutated by (+/-)-anti-BPDE was essentially random in umethylated and methylated p53, suggesting that 5'-CpG-3' islands did not direct mutations in the assay. These data suggest that smoking may cause mutations in p53 by formation of PAH o-quinones, which produce reactive oxygen species. The resultant 8-oxo-dGuo yields a pattern of mutations but not a spectrum consistent with that seen in lung cancer; we suggest that the emergence of the spectrum requires biological selection.