State Key Laboratory of Pharmaceutical Biotechnology, Department of Gastroenterology, Drum Tower Hospital, The Affiliated Hospital of Nanjing University Medical School, Nanjing, Jiangsu 210093, China.
Department of Cardiothoracic Surgery, Nanjing Drum Tower Hospital, Medical School of Nanjing University, Nanjing, China.
Nucleic Acids Res. 2020 Jul 27;48(13):7027-7040. doi: 10.1093/nar/gkaa504.
Methylation of miRNAs at the 2'-hydroxyl group on the ribose at 3'-end (2'-O-methylation, 2'Ome) is critical for miRNA function in plants and Drosophila. Whether this methylation phenomenon exists for mammalian miRNA remains unknown. Through LC-MS/MS analysis, we discover that majority of miR-21-5p isolated from human non-small cell lung cancer (NSCLC) tissue possesses 3'-terminal 2'Ome. Predominant 3'-terminal 2'Ome of miR-21-5p in cancer tissue is confirmed by qRT-PCR and northern blot after oxidation/β-elimination procedure. Cancerous and the paired non-cancerous lung tissue miRNAs display different pattern of 3'-terminal 2'Ome. We further identify HENMT1 as the methyltransferase responsible for 3'-terminal 2'Ome of mammalian miRNAs. Compared to non-methylated miR-21-5p, methylated miR-21-5p is more resistant to digestion by 3'→5' exoribonuclease polyribonucleotide nucleotidyltransferase 1 (PNPT1) and has higher affinity to Argonaute-2, which may contribute to its higher stability and stronger inhibition on programmed cell death protein 4 (PDCD4) translation, respectively. Our findings reveal HENMT1-mediated 3'-terminal 2'Ome of mammalian miRNAs and highlight its role in enhancing miRNA's stability and function.
miRNA 在核糖 3' 端 2' 羟基上的甲基化（2'-O-甲基化，2'Ome）对于植物和果蝇中的 miRNA 功能至关重要。这种甲基化现象是否存在于哺乳动物 miRNA 中尚不清楚。通过 LC-MS/MS 分析，我们发现从人非小细胞肺癌（NSCLC）组织中分离的大多数 miR-21-5p 都具有 3' 末端 2'Ome。通过氧化/β-消除处理后，qRT-PCR 和 northern blot 证实了癌症组织中 miR-21-5p 的主要 3' 末端 2'Ome。癌症组织和配对的非癌组织 miRNA 显示出不同的 3' 末端 2'Ome 模式。我们进一步鉴定出 HENMT1 是负责哺乳动物 miRNA 3' 末端 2'Ome 的甲基转移酶。与未甲基化的 miR-21-5p 相比，甲基化的 miR-21-5p 对 3'→5' 外切核糖核酸酶多核苷酸核苷酸转移酶 1（PNPT1）的消化更具抗性，并且与 Argonaute-2 的亲和力更高，这可能分别有助于其更高的稳定性和对程序性细胞死亡蛋白 4（PDCD4）翻译的更强抑制作用。我们的发现揭示了 HENMT1 介导的哺乳动物 miRNA 的 3' 末端 2'Ome，并强调了其在增强 miRNA 的稳定性和功能中的作用。
