Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, Massachusetts, United States of America.
PLoS Genet. 2010 Dec 16;6(12):e1001247. doi: 10.1371/journal.pgen.1001247.
Decades of study have revealed more than 100 ribonucleoside structures incorporated as post-transcriptional modifications mainly in tRNA and rRNA, yet the larger functional dynamics of this conserved system are unclear. To this end, we developed a highly precise mass spectrometric method to quantify tRNA modifications in Saccharomyces cerevisiae. Our approach revealed several novel biosynthetic pathways for RNA modifications and led to the discovery of signature changes in the spectrum of tRNA modifications in the damage response to mechanistically different toxicants. This is illustrated with the RNA modifications Cm, m(5)C, and m(2) (2)G, which increase following hydrogen peroxide exposure but decrease or are unaffected by exposure to methylmethane sulfonate, arsenite, and hypochlorite. Cytotoxic hypersensitivity to hydrogen peroxide is conferred by loss of enzymes catalyzing the formation of Cm, m(5)C, and m(2) (2)G, which demonstrates that tRNA modifications are critical features of the cellular stress response. The results of our study support a general model of dynamic control of tRNA modifications in cellular response pathways and add to the growing repertoire of mechanisms controlling translational responses in cells.
数十年的研究揭示了超过 100 种核糖核苷结构，它们主要作为转录后修饰被整合到 tRNA 和 rRNA 中，但这个保守系统的更大功能动态尚不清楚。为此，我们开发了一种高度精确的质谱分析方法来定量测定酿酒酵母中的 tRNA 修饰。我们的方法揭示了几种新的 RNA 修饰生物合成途径，并导致在对不同机制的有毒物质的损伤反应中发现了 tRNA 修饰谱的特征变化。这可以用 RNA 修饰 Cm、m(5)C 和 m(2)（2）G 来说明，它们在过氧化氢暴露后增加，但在用甲基甲烷磺酸酯、亚砷酸盐和次氯酸盐暴露时减少或不受影响。过氧化氢引起的细胞毒性超敏性是由催化 Cm、m(5)C 和 m(2)（2）G 形成的酶缺失引起的，这表明 tRNA 修饰是细胞应激反应的关键特征。我们研究的结果支持了 tRNA 修饰在细胞反应途径中动态控制的一般模型，并为控制细胞中翻译反应的机制不断增加提供了补充。
