Transcriptome-wide mapping of 5-methylcytidine RNA modifications in bacteria, archaea, and yeast reveals m5C within archaeal mRNAs.
Transcriptome-wide mapping of 5-methylcytidine RNA modifications in bacteria, archaea, and yeast reveals m5C within archaeal mRNAs.
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DOI：
10.1371/journal.pgen.1003602
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发表时间：
2013-06
期刊：
影响因子：
4.5
通讯作者：
Sorek R
中科院分区：
文献类型：
--
作者：
Edelheit S;Schwartz S;Mumbach MR;Wurtzel O;Sorek R
40多年前，在多种生物的tRNA和rRNA分子中首次观察到5-甲基胞苷（m5 C）的存在。然而，由于使用低通量方法，这种修饰的检测仅限于特定的、丰富的RNA种类。为了获得跨越生命的三个领域的m5 C的高分辨率、系统性和全面的转录组范围的概述，我们对来自革兰氏阳性（B. subtilis）和革兰氏阴性（E. coli）细菌、古细菌（S. solfataricus）和真核生物（S. cerevisiae），然后进行大规模平行测序。我们能够恢复大多数以前记录的m5 C位点的rRNA在四种生物体，并确定了几个新的网站在酵母和古细菌的rRNA。我们的分析还允许在酵母和古细菌中的64个tRNA中的甲基化m5 C位置的定量，揭示了这些生物体的甲基化模式之间的化学计量差异。还发现了不存在m5 C的tRNA分子。有趣的是，我们检测到古细菌mRNA中的m5 C位点，并确定了指导S.太阳神我们的研究结果，这是使用m5 C特异性RNA免疫沉淀验证，提供了第一个证据的古细菌中的mRNA修饰，这表明这种模式的转录后调控延伸到真核生物域之外。核糖核酸普遍用于以基因转录物的形式表达遗传信息。虽然我们设想RNA仅仅是DNA四碱基密码的拷贝，但特定RNA碱基的修饰可以扩展信息密码。这种修饰在转运RNA（tRNA）和核糖体RNA（rRNA）中大量存在，它们有助于翻译保真度和核糖体组装。最近在真核生物中的研究表明，mRNA修饰，如RNA编辑（将腺苷碱基转化为肌苷），N6-腺嘌呤甲基化（m6 A）和5-甲基胞苷（m5 C）可以改变编码序列，改变剪接模式或改变RNA稳定性。然而，迄今为止还没有记录细菌或古细菌中的mRNA修饰。我们已经使用了一种方法，该方法能够在给定生物体中的所有表达基因中映射m5 C修饰。将这种方法应用于模型细菌，古细菌和真菌微生物，使我们能够揭示这些生物体中修饰的RNA碱基，并提供这些修饰的准确和灵敏的图谱。在古细菌中，我们记录了多个基因的mRNA受到RNA修饰，这表明类似于真核生物，这些生物可能利用mRNA修饰作为基因调控的机制。
The presence of 5-methylcytidine (m5C) in tRNA and rRNA molecules of a wide variety of organisms was first observed more than 40 years ago. However, detection of this modification was limited to specific, abundant, RNA species, due to the usage of low-throughput methods. To obtain a high resolution, systematic, and comprehensive transcriptome-wide overview of m5C across the three domains of life, we used bisulfite treatment on total RNA from both gram positive (B. subtilis) and gram negative (E. coli) bacteria, an archaeon (S. solfataricus) and a eukaryote (S. cerevisiae), followed by massively parallel sequencing. We were able to recover most previously documented m5C sites on rRNA in the four organisms, and identified several novel sites in yeast and archaeal rRNAs. Our analyses also allowed quantification of methylated m5C positions in 64 tRNAs in yeast and archaea, revealing stoichiometric differences between the methylation patterns of these organisms. Molecules of tRNAs in which m5C was absent were also discovered. Intriguingly, we detected m5C sites within archaeal mRNAs, and identified a consensus motif of AUCGANGU that directs methylation in S. solfataricus. Our results, which were validated using m5C-specific RNA immunoprecipitation, provide the first evidence for mRNA modifications in archaea, suggesting that this mode of post-transcriptional regulation extends beyond the eukaryotic domain. Ribonucleic acids are universally used to express genetic information in the form of gene transcripts. Although we envision RNA as a mere copy of the DNA four-base code, modification of specific RNA bases can expand the information code. Such modifications are abundant in transfer RNA (tRNA) and ribosomal RNA (rRNA), where they contribute to translation fidelity and ribosome assembly. Recent studies in eukaryotes have shown that mRNA modifications such as RNA-editing (conversion of an adenosine base to inosine), N6-adenine methylation (m6A), and 5-methylcytidine (m5C) can change the coding sequence, alter splicing patterns, or change RNA stability. However, no mRNA modifications in bacteria or archaea have been documented to date. We have used an approach that enables mapping of the m5C modifications across all expressed genes in a given organism. Applying this approach on model bacterial, archaeal, and fungal microorganisms enabled us to reveal the modified RNA bases in these organisms, and to provide an accurate and sensitive map of these modifications. In archaea, we documented multiple genes whose mRNAs are subject to RNA modification, suggesting that similar to eukaryotes, these organisms may utilize mRNA modifications as a mechanism for gene regulation.
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