Translational regulation during cigarette smoking-induced reprogramming of the tRNA epitranscriptome, in vitro and in a mouse smoking model

1.项目摘要 吸烟会导致心血管疾病、慢性阻塞性肺疾病（COPD）和肺部疾病。 癌暴露于环境烟草烟雾（ETS）引起的疾病的病因学受到以下因素的混淆： ETS的复杂性，包括直接作用和代谢活化的> 4000种化学物质的混合物 有毒物质虽然所有生物体都通过调节基因表达来应对环境暴露，但我们所知甚少。 关于将毒物暴露与细胞生存和疾病联系起来的翻译机制。在这里，我们寻求 了解几十种修饰的核糖核苷在tRNA中的作用-tRNA表转录组-在 调节细胞对ETS暴露的反应。使用独特的计算和分析平台，我们有 表明（1）毒物暴露导致酵母和哺乳动物表位转录组的特征变化， 细胞，和（2）毒物诱导的tRNA表位转录组重编程调节蛋白质水平， 促进来自酵母中应激反应基因家族的密码子偏向mRNA的选择性翻译。我们 在细菌、酵母和哺乳动物细胞中的研究1 - 7也表明，关键的表转录组书写者的缺陷 由于应激反应蛋白的翻译受损，酶使细胞对特定毒物的杀伤敏感。 我们假设暴露于ETS的复杂混合物中会重新编程tRNA表位转录组 以反映ETS中的主要化学应激源，并且改变的RNA修饰将调节 细胞行为通过选择性翻译密码子偏向的应激反应mRNAs。为了支持这一点，我们有 在暴露于药物和毒物的大鼠肝脏中观察到试剂特异性表位转录组重编程 (e.g.,砷; NTP DrugMatrix），以及烷基化和氧化特异性标志tRNA修饰的变化， 酵母和人类细胞。我们的表位转录组书写缺陷（Alkbh 8-null）小鼠显示tRNA修饰 翻译氧化解毒硒蛋白所需的系统对于暴露于 萘，一种P450活化的、产生氧化剂的多环芳烃（PAH）。三个目标 从体外培养的小鼠细胞研究到小鼠ETS暴露模型，我们测试了 表位转录组和翻译调节在细胞对ETS的应答中起重要作用。 !

1. PROJECT SUMMARY Cigarette smoking causes cardiovascular disease, chronic obstructive pulmonary disease (COPD), and lung cancer. The etiology of diseases caused by exposure to environmental tobacco smoke (ETS) is confounded by the complexity of ETS as a mixture of >4000 chemicals, including direct-acting and metabolically-activated toxicants. While all organisms respond to environmental exposures by regulating gene expression, we know little about the translational mechanisms linking toxicant exposure to cell survival and disease. Here we seek to understand the role of the dozens of modified ribonucleosides in tRNA – the tRNA epitranscriptome – in regulating the cell response to ETS exposure. Using a unique computational and analytical platform, we have shown that (1) toxicant exposures cause signature changes in the epitranscriptome of yeast and mammalian cells, and (2) that toxicant-induced reprogramming of the tRNA epitranscriptome regulates protein levels by promoting the selective translation of codon-biased mRNAs from families of stress-response genes in yeast. Our studies in bacteria, yeast and mammalian cells1-7 also show that deficiencies in key epitranscriptomic writer enzymes sensitize cells to killing by specific toxicants due to corrupted translation of stress response proteins. We hypothesize that exposure to the complex mixture of ETS will reprogram the tRNA epitranscriptome to reflect the predominant chemical stressors in ETS and that the altered RNA modifications will regulate cell behavior by selective translation of codon-biased stress response mRNAs. In support of this, we have observed agent-specific epitranscriptome reprogramming in the liver from rats exposed to drugs and toxicants (e.g., arsenic; NTP DrugMatrix), and alkylation- and oxidation-specific signature tRNA modification changes in yeast and human cells. Our epitranscriptomic writer-deficient (Alkbh8-null) mice showed that tRNA modification systems required for translating oxidant-detoxifying selenoproteins are vital to surviving exposure to naphthalene, a P450-activated, oxidant-generating, polycyclic aromatic hydrocarbon (PAH) in ETS. In three aims ranging from in vitro studies in cultured mouse cells to a mouse ETS exposure model, we test the idea that the epitranscriptome and translational regulation play an important role in the cell response to ETS. !