PDF(Pubmed)
Abstract：
Recoding viral genomes by introducing numerous synonymous but suboptimal codon pairs-called codon-pair deoptimization (CPD)-provides new types of live-attenuated vaccine candidates. The large number of nucleotide changes resulting from CPD should provide genetic stability to the attenuating phenotype, but this has not been rigorously tested. Human respiratory syncytial virus in which the G and F surface glycoprotein ORFs were CPD (called Min B) was temperature-sensitive and highly restricted in vitro. When subjected to selective pressure by serial passage at increasing temperatures, Min B substantially regained expression of F and replication fitness. Whole-genome deep sequencing showed many point mutations scattered across the genome, including one combination of six linked point mutations. However, their reintroduction into Min B provided minimal rescue. Further analysis revealed viral genomes bearing very large internal deletions (LD genomes) that accumulated after only a few passages. The deletions relocated the CPD F gene to the first or second promoter-proximal gene position. LD genomes amplified de novo in Min B-infected cells were encapsidated, expressed high levels of F, and complemented Min B replication in trans This study provides insight on a variation of the adaptability of a debilitated negative-strand RNA virus, namely the generation of defective minihelper viruses to overcome its restriction. This is in contrast to the common \"defective interfering particles\" that interfere with the replication of the virus from which they originated. To our knowledge, defective genomes that promote rather than inhibit replication have not been reported before in RNA viruses.
摘要：
通过引入许多同义但次优的密码子对(称为密码子对去优化(CPD))来重新编码病毒基因组提供了新型的减毒活疫苗候选物。CPD导致的大量核苷酸变化应该为减毒表型提供遗传稳定性。但是这还没有经过严格的测试。其中G和F表面糖蛋白ORFs为CPD的人呼吸道合胞病毒（称为MinB）对温度敏感且在体外具有高度限制性。当在升高的温度下通过串联通道承受选择性压力时，MinB基本上恢复了F和复制适合度的表达。全基因组深度测序显示许多点突变分散在基因组中，包括六个连锁点突变的组合。然而,他们重新引入MinB提供了最小的救援。进一步的分析显示病毒基因组带有非常大的内部缺失（LD基因组），仅在几次传代后积累。缺失将CPDF基因重新定位到第一或第二启动子近端基因位置。在MinB感染细胞中从头扩增的LD基因组被衣壳化，表示高水平的F，并补充了MinB的反式复制本研究提供了对衰弱的负链RNA病毒适应性变化的见解，即产生有缺陷的微型辅助病毒以克服其限制。这与常见的“有缺陷的干扰颗粒”相反，后者会干扰它们起源的病毒的复制。据我们所知,以前在RNA病毒中没有报道过促进而不是抑制复制的缺陷基因组。
