Internal RNAs overlapping coding sequences can drive the production of alternative proteins in archaea

Prokaryotic genomes show a high level of information compaction often with different molecules transcribed from the same <i>locus</i>. Although antisense RNAs have been relatively well studied, RNAs in the same strand, internal RNAs (intraRNAs), are still poorly understood. The question of how common is the translation of overlapping reading frames remains open. We address this question in the model archaeon <i>Halobacterium salinarum</i>. In the present work we used differential RNA-seq (dRNA-seq) in <i>H. salinarum</i> NRC-1 to locate intraRNA signals in subsets of internal transcription start sites (iTSS) and establish the open reading frames associated to them (intraORFs). Using C-terminally flagged proteins, we experimentally observed isoforms accurately predicted by intraRNA translation for <i>kef1, acs3</i> and <i>orc4</i> genes. We also recovered from the literature and mass spectrometry databases several instances of protein isoforms consistent with intraRNA translation such as the gas vesicle protein gene <i>gvpC1</i>. We found evidence for intraRNAs in horizontally transferred genes such as the chaperone <i>dnaK</i> and the aerobic respiration related <i>cydA</i> in both <i>H. salinarum</i> and <i>Escherichia coli</i>. Also, intraRNA translation evidence in <i>H. salinarum, E. coli</i> and yeast of a universal elongation factor (<i>aEF-2, fusA</i> and <i>eEF</i>-2) suggests that this is an ancient phenomenon present in all domains of life.

原核生物基因组具有高度的信息压缩特性，同一基因座（locus）往往可转录产生多种不同的RNA分子。尽管反义RNA（antisense RNAs）的研究已较为深入，但同链RNA中的内部RNA（intraRNAs）仍有待进一步探索。关于重叠阅读框的翻译究竟有多普遍，这一科学问题尚未得到明确解答。本研究以模式古菌盐生盐杆菌（Halobacterium salinarum）为研究体系，对该问题展开探究。本研究利用差异RNA测序（differential RNA-seq, dRNA-seq）技术，对盐生盐杆菌NRC-1（H. salinarum NRC-1）进行转录组分析，在内部转录起始位点（internal transcription start sites, iTSS）的子集区域中定位内部RNA信号，并确定与之对应的内部开放阅读框（intraORFs）。研究人员通过C端标记蛋白实验，成功观测到kef1、acs3及orc4基因经内部RNA翻译所精准预测得到的蛋白同工型。此外，本研究从已发表文献及质谱数据库中挖掘到多例与内部RNA翻译相符的蛋白同工型实例，例如气体囊泡蛋白基因gvpC1。本研究在盐生盐杆菌与大肠杆菌（Escherichia coli）的水平转移基因中，均发现了内部RNA存在的证据，例如分子伴侣基因dnaK以及参与有氧呼吸的cydA基因。此外，在盐生盐杆菌、大肠杆菌及酵母中，针对通用延伸因子（universal elongation factor, aEF-2、fusA及eEF-2）的内部RNA翻译证据表明，这一现象是一种古老的生物学机制，广泛存在于所有生命域（domains of life）中。