核糖体图谱技术在植物学研究中的应用

doi:10.11983/CBB22028

摘要/Abstract

摘要： 随着高通量测序技术的持续发展和进步, 开发出很多新颖的测序技术, 为诸多悬而未决的生物学难题提供了解决方案。其中, 核糖体图谱技术能够在全基因组水平和单核苷酸分辨率上监控细胞内的翻译事件, 填补了转录组学和蛋白质组学研究之间的空隙。核糖体图谱技术不仅能够鉴定处于翻译状态的RNA分子, 还能够精确定位RNA分子上正在翻译的核苷酸, 进而准确描绘RNA分子上的开放阅读框。此外, 结合转录组测序数据, 核糖体图谱技术还可以确定每个转录本上的核糖体数量, 从而计算每个转录本的翻译效率。目前, 核糖体图谱技术已成功应用于动物、植物和微生物等研究领域, 加深了人们对翻译调控机制的认识。然而, 由于植物细胞和组织的特性, 核糖体图谱技术在植物学研究中的应用仍然存在局限。该文综述了核糖体图谱技术的实验原理, 以及在植物学研究中的相关进展。

关键词: 核糖体图谱, 翻译调控, 上游开放阅读框

Abstract: With the great development and progress of high-throughput sequencing (HTS) technologies, many newborn techniques have provided solutions to unresolved biological puzzles. Ribosome profiling has emerged as an HTS-based technique that facilitates the genome-wide investigation of translation at single-nucleotide resolution, and it resolves the missing link between transcriptome and translatome. Ribosome profiling not only identifies the mRNAs in translation, but also provides precise positions of the ribosomes on mRNA to help identify the coding regions of the mRNAs. In addition, combined with the parallel RNA sequencing, ribosome profiling can determine the translation efficiency for each mRNA by comparing the rates of protein synthesis and the abundance of mRNAs. Based on its extraordinary advances, ribosome profiling has been successfully applied in animals, plants and microbes, and has thoroughly enhanced our understanding of translational regulation. However, due to the restrictions caused by the physical characteristics of plant cells and tissues, there are still considerable limitations in the application of ribosome profiling in plants. Here, we introduce the experimental principle of ribosome profiling and review the recent progresses made in botanical researches.

Key words: ribosome profiling, translational regulation, upstream open reading frame

王豫颖, 王威浩. 核糖体图谱技术在植物学研究中的应用. 植物学报, 2022, 57(5): 673-683.

Wang Yuying, Wang Weihao. Recent Advances of Ribosome Profiling in Plants. Chinese Bulletin of Botany, 2022, 57(5): 673-683.

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链接本文: https://www.chinbullbotany.com/CN/10.11983/CBB22028

https://www.chinbullbotany.com/CN/Y2022/V57/I5/673

图/表 1

图1 核糖体图谱实验流程示意图 (A) 在核糖体图谱技术(Ribo-seq)中, 多聚核糖体经RNA酶消化成单核糖体, 约30 nt的受核糖体保护的mRNA片段(RPFs)未被RNA酶降解; 将单核糖体解离, 并从中分离和纯化RPFs用于高通量测序, 仅mRNA的蛋白编码区有测序信号; (B) 搭配转录组测序(RNA-seq), 可计算出每条转录本的翻译效率, 在转录组测序中整个转录本上均有测序信号。5′UTR: 5′端非翻译区; 3′UTR: 3′端非翻译区; ORF: 开放阅读框

Figure 1 A schematic diagram of ribosome profiling (A) In the experimental procedure of ribosome profiling (Ribo-seq), polysomes are degraded into monosomes by RNase, resulting in the preservation of the ~30 nucleotides of the ribosome-protected fragments (RPFs) owing to the resistance of the ribosome to the digestion. After isolation and purification from the monosomes, the RPFs are further used for high-throughput sequencing. The ribosome footprints typically show precise positioning within the coding sequence of a mRNA; (B) Ribosome profiling is usually accompanied by a parallel RNA sequencing (RNA-seq) to calculate the translation efficiency of the transcripts. RNA-seq captures mRNA fragments covering the entire transcripts. 5′UTR: 5′ untranslated region; 3′UTR: 3′ untranslated region; ORF: Open reading frame

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