Department of Chemistry, Department of Biochemistry and Molecular Biology, and Institute for Biophysical Dynamics, The University of Chicago, Chicago, IL 60637, USA; Howard Hughes Medical Institute, The University of Chicago, Chicago, IL 60637, USA.
Liver Cancer Institute, Zhongshan Hospital, Fudan University, Shanghai 200032, China; Institute of Biomedical Sciences, Fudan University, Shanghai 200032, China.
Mol Cell. 2019 Jun 20;74(6):1304-1316.e8. doi: 10.1016/j.molcel.2019.03.036. Epub 2019 Apr 25.
N-methylguanosine (mG) is a positively charged, essential modification at the 5' cap of eukaryotic mRNA, regulating mRNA export, translation, and splicing. mG also occurs internally within tRNA and rRNA, but its existence and distribution within eukaryotic mRNA remain to be investigated. Here, we show the presence of internal mG sites within mammalian mRNA. We then performed transcriptome-wide profiling of internal mG methylome using mG-MeRIP sequencing (MeRIP-seq). To map this modification at base resolution, we developed a chemical-assisted sequencing approach that selectively converts internal mG sites into abasic sites, inducing misincorporation at these sites during reverse transcription. This base-resolution mG-seq enabled transcriptome-wide mapping of mG in human tRNA and mRNA, revealing distribution features of the internal mG methylome in human cells. We also identified METTL1 as a methyltransferase that installs a subset of mG within mRNA and showed that internal mG methylation could affect mRNA translation.
N6-甲基鸟嘌呤（mG）是真核 mRNA 5'帽的一种带正电荷的必需修饰，调节 mRNA 的输出、翻译和剪接。mG 也存在于 tRNA 和 rRNA 内部，但它在真核 mRNA 中的存在和分布仍有待研究。在这里，我们展示了哺乳动物 mRNA 内部 mG 位点的存在。然后，我们使用 mG-MeRIP 测序（MeRIP-seq）进行了全转录组范围的内部 mG 甲基组谱分析。为了在碱基分辨率上绘制这种修饰，我们开发了一种化学辅助测序方法，该方法选择性地将内部 mG 位点转化为无碱基位点，在反转录过程中导致这些位点的错误掺入。这种碱基分辨率的 mG-seq 能够在人类 tRNA 和 mRNA 中进行全转录组范围的 mG 作图，揭示了人类细胞中内部 mG 甲基组的分布特征。我们还鉴定出 METTL1 是一种将一部分 mG 安装在 mRNA 中的甲基转移酶，并表明内部 mG 甲基化可能影响 mRNA 翻译。
