基于质谱的tRNA从头直接测序和多RNA修饰的定量定

本期文章：《美国化学会志》：Online/在线发表

美国纽约理工学院Shenglong Zhang团队取得一项新突破。他们的最新研究提出了基于质谱的tRNA从头直接测序和多RNA修饰的定量定位。2024年9月4日，国际知名学术期刊《美国化学会杂志》发表了这一成果。

尽管RNA的下一代测序(NGS)已被广泛的应用，但同时对多个RNA核苷酸修饰进行直接测序和定量定位仍然具有挑战性。基于质谱(MS)的测序可以直接对所有RNA修饰进行测序，而不局限于特定的修饰，但它需要一个完美的MS梯子，但很少有tRNA可以提供这个完美的梯子。

在这项研究，课题组描述了一种MS梯子互补测序方法(MLC-Seq)，绕过了完美的梯子的要求，允许以单核苷酸精度对具有多个核苷酸修饰的，全长异质细胞tRNA进行从头MS测序。

与NGS方法不同，MLC-Seq保留了RNA序列多样性和修饰信息，揭示了新的详细的化学计量tRNA修饰谱及其在脱烷基酶AlkB处理后的变化。它也可以与参考序列相结合，提供各种tRNA和总tRNA样品中修饰的定量分析。MLC-Seq能够系统、定量、位点特异性地绘制RNA修饰图谱，揭示tRNA真正完整的信息内容。

附：英文原文

Title: Mass Spectrometry-Based Direct Sequencing of tRNAs De Novo and Quantitative Mapping of Multiple RNA Modifications

Author: Xiaohong Yuan, Yue Su, Benjamin Johnson, Michele Kirchner, Xudong Zhang, Sihang Xu, Sophia Jiang, Jing Wu, Shundi Shi, James J. Russo, Qi Chen, Shenglong Zhang

Issue&Volume: September 4, 2024

Abstract: Despite the extensive use of next-generation sequencing (NGS) of RNA, simultaneous direct sequencing and quantitative mapping of multiple RNA nucleotide modifications remains challenging. Mass spectrometry (MS)-based sequencing can directly sequence all RNA modifications without being limited to specific ones, but it requires a perfect MS ladder that few tRNAs can provide. Here, we describe an MS ladder complementation sequencing approach (MLC-Seq) that circumvents the perfect ladder requirement, allowing de novo MS sequencing of full-length heterogeneous cellular tRNAs with multiple nucleotide modifications at single-nucleotide precision. Unlike NGS-based methods, which lose RNA modification information, MLC-Seq preserves RNA sequence diversity and modification information, revealing new detailed stoichiometric tRNA modification profiles and their changes upon treatment with the dealkylating enzyme AlkB. It can also be combined with reference sequences to provide quantitative analysis of diverse tRNAs and modifications in total tRNA samples. MLC-Seq enables systematic, quantitative, and site-specific mapping of RNA modifications, revealing the truly complete informational content of tRNA.

DOI: 10.1021/jacs.4c07280

Source: https://pubs.acs.org/doi/full/10.1021/jacs.4c07280

期刊信息

JACS：《美国化学会志》，创刊于1879年。隶属于美国化学会，最新IF：16.383
官方网址：https://pubs.acs.org/journal/jacsat
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