Infection and Immunity Division, Roslin Institute, University of Edinburgh, Easter Bush, Edinburgh EH25 9RG, UK and School of Life Sciences, University of Warwick, Coventry CV4 7AL, UK.
Nucleic Acids Res. 2014 Apr;42(7):4527-45. doi: 10.1093/nar/gku075. Epub 2014 Jan 26.
Most RNA viruses infecting mammals and other vertebrates show profound suppression of CpG and UpA dinucleotide frequencies. To investigate this functionally, mutants of the picornavirus, echovirus 7 (E7), were constructed with altered CpG and UpA compositions in two 1.1-1.3 Kbase regions. Those with increased frequencies of CpG and UpA showed impaired replication kinetics and higher RNA/infectivity ratios compared with wild-type virus. Remarkably, mutants with CpGs and UpAs removed showed enhanced replication, larger plaques and rapidly outcompeted wild-type virus on co-infections. Luciferase-expressing E7 sub-genomic replicons with CpGs and UpAs removed from the reporter gene showed 100-fold greater luminescence. E7 and mutants were equivalently sensitive to exogenously added interferon-β, showed no evidence for differential recognition by ADAR1 or pattern recognition receptors RIG-I, MDA5 or PKR. However, kinase inhibitors roscovitine and C16 partially or entirely reversed the attenuated phenotype of high CpG and UpA mutants, potentially through inhibition of currently uncharacterized pattern recognition receptors that respond to RNA composition. Generating viruses with enhanced replication kinetics has applications in vaccine production and reporter gene construction. More fundamentally, the findings introduce a new evolutionary paradigm where dinucleotide composition of viral genomes is subjected to selection pressures independently of coding capacity and profoundly influences host-pathogen interactions.
大多数感染哺乳动物和其他脊椎动物的 RNA 病毒表现出对 CpG 和 UpA 二核苷酸频率的明显抑制。为了研究其功能，构建了小核糖核酸病毒（肠道病毒 7，E7）的突变体，在两个 1.1-1.3 Kb 区域改变了 CpG 和 UpA 的组成。与野生型病毒相比，CpG 和 UpA 频率增加的突变体表现出复制动力学受损和更高的 RNA/感染性比率。值得注意的是，去除 CpG 和 UpA 的突变体显示出增强的复制能力，更大的斑块，并且在共感染时迅速竞争超过野生型病毒。从报告基因中去除 CpG 和 UpA 的含有 Luciferase 的 E7 亚基因组复制子显示出 100 倍的发光强度。E7 和突变体对外源性添加的干扰素-β具有同等的敏感性，没有证据表明 ADAR1 或模式识别受体 RIG-I、MDA5 或 PKR 对它们有差异识别。然而，激酶抑制剂罗司维亭和 C16 部分或完全逆转了高 CpG 和 UpA 突变体的减弱表型，可能是通过抑制目前未知的对 RNA 组成有反应的模式识别受体。产生具有增强复制动力学的病毒在疫苗生产和报告基因构建方面具有应用价值。更根本的是，这些发现引入了一个新的进化范例，即病毒基因组的二核苷酸组成独立于编码能力而受到选择压力的影响，并深刻地影响了宿主-病原体相互作用。
