Positive-sense single-stranded RNA viruses form the largest and most diverse group of eukaryote-infecting viruses. Their genomes comprise one or more segments of coding-sense RNA that function directly as messenger RNAs upon release into the cytoplasm of infected cells. Positive-sense RNA viruses are generally accepted to encode proteins solely on the positive strand. However, we previously identified a surprisingly long (∼1,000-codon) open reading frame (ORF) on the negative strand of some members of the family Narnaviridae which, together with RNA bacteriophages of the family Leviviridae, form a sister group to all other positive-sense RNA viruses. Here, we completed the genomes of three mosquito-associated narnaviruses, all of which have the long reverse-frame ORF. We systematically identified narnaviral sequences in public data sets from a wide range of sources, including arthropod, fungal, and plant transcriptomic data sets. Long reverse-frame ORFs are widespread in one clade of narnaviruses, where they frequently occupy >95 per cent of the genome. The reverse-frame ORFs correspond to a specific avoidance of CUA, UUA, and UCA codons (i.e. stop codon reverse complements) in the forward-frame RNA-dependent RNA polymerase ORF. However, absence of these codons cannot be explained by other factors such as inability to decode these codons or GC3 bias. Together with other analyses, we provide the strongest evidence yet of coding capacity on the negative strand of a positive-sense RNA virus. As these ORFs comprise some of the longest known overlapping genes, their study may be of broad relevance to understanding overlapping gene evolution and de novo origin of genes.

中文翻译：

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一组正义RNA病毒中负链编码序列的情况。

正链单链RNA病毒是感染真核生物的最大和最多样化的组。它们的基因组包含一个或多个编码有义RNA片段，这些片段在释放到感染细胞的细胞质中后直接充当信使RNA。通常公认的正义RNA病毒仅在正链上编码蛋白质。然而，我们先前在纳纳病毒科某些成员的负链上鉴定到一个令人惊讶的长阅读框（〜1,000个密码子），它与左旋病毒科的RNA噬菌体一起构成了其他所有阳性细胞的姐妹组。 -有义RNA病毒。在这里，我们完成了三种与蚊子相关的纳纳病毒的基因组，它们均具有长的反向框架ORF。我们从包括节肢动物，真菌和植物转录组数据集在内的各种来源中，系统地识别了公共数据集中的纳那病毒序列。较长的反向框架ORF广泛分布于一组纳纳病毒中，它们经常占据基因组的95％以上。反向框ORF对应于前向框依赖RNA的RNA聚合酶ORF中对CUA，UUA和UCA密码子（即终止密码子反向补体）的特定回避。但是，这些密码子的缺失无法用其他因素来解释，例如无法解码这些密码子或GC3偏倚。连同其他分析，我们提供了迄今为止最有力的证据，证明了在正义RNA病毒负链上的编码能力。由于这些ORF包含一些已知最长的重叠基因，