Inactivation of Genes by Frameshift Mutations Provides Rapid Adaptation of an Attenuated Vaccinia Virus.
Inactivation of Genes by Frameshift Mutations Provides Rapid Adaptation of an Attenuated Vaccinia Virus.
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通过移码突变使基因失活可以使减毒痘苗病毒快速适应。
DOI：
10.1128/jvi.01053-20
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发表时间：
2020
影响因子：
5.4
通讯作者：
Moss,Bernard
中科院分区：
文献类型：
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作者：
Senkevich,TatianaG;Zhivkoplias,ErikK;Weisberg,AndreaS;Moss,Bernard
与 RNA 病毒不同，大多数 DNA 病毒使用高保真聚合酶复制其基因组，很少出现碱基替换错误。然而，实验进化研究表明，在减毒痘苗病毒（VACV）连续传代过程中，适应性突变会快速获得。适应发生的一种方式是通过手风琴机制，其中基因拷贝数增加，随后进行碱基替换，最后基因拷贝数收缩。在这里，我们展示了在减毒VACV传代过程中，由基因失活移码机制介导的多种适应性突变的快速获得。通过将 VACV A8R 中间转录因子基因与粘液瘤病毒直向同源物交换来实现减毒。减毒病毒的三个平行传代中总共发生了六个不同基因的七个突变。最常见的突变是 5 到 7 个 As 或 T 运行中的单核苷酸插入或删除，尽管也发生了 11 个核苷酸的删除，导致移码和过早终止密码子。在 10 轮传代中，减毒的 VACV 被突变病毒所取代。实验结束时，几乎所有剩余的病毒都有一种固定突变和一种或多种额外突变。尽管转录装置中的核苷酸取代导致了两个低频突变，但编码成熟病毒体蛋白质成分的基因（即 A26L、G6R 和 A14.5L）的移码实现了 74% 至 98% 的固定。通过制作删除 A26L 或 G6R 或两者的重组 VACV，证实了突变的适应性作用，这增加了病毒复制水平并降低了颗粒/PFU 比率。重要性基因失活被认为是正痘病毒进化的重要驱动因素。尽管牛痘病毒含有每个正痘病毒物种中存在的基因的完整直系同源物，但该属的所有其他成员中的许多基因都失活了。正痘病毒在细胞培养物中广泛传代时，可能会发生其他基因的失活，导致体内减毒，这是一种制造疫苗的策略。在大多数情况下，多个病毒基因的失活是否会增强宿主细胞中的复制或具有中性作用尚不清楚。使用涉及减毒痘苗病毒连续传代的实验进化方案，快速获得失活移码突变。仅经过 10 轮传代后，起始的减毒痘苗病毒就被具有一种固定突变和一种或多种额外突变的病毒所取代。实验进化过程中多种失活突变的高频率模拟了它们在正常进化和广泛病毒传代以制造疫苗株期间的获得。
Unlike RNA viruses, most DNA viruses replicate their genomes with high-fidelity polymerases that rarely make base substitution errors. Nevertheless, experimental evolution studies have revealed rapid acquisition of adaptive mutations during serial passage of attenuated vaccinia virus (VACV). One way in which adaptation can occur is by an accordion mechanism in which the gene copy number increases followed by base substitutions and, finally, contraction of the gene copy number. Here, we show rapid acquisition of multiple adaptive mutations mediated by a gene-inactivating frameshift mechanism during passage of an attenuated VACV. Attenuation had been achieved by exchanging the VACV A8R intermediate transcription factor gene with the myxoma virus ortholog. A total of seven mutations in six different genes occurred in three parallel passages of the attenuated virus. The most frequent mutations were single-nucleotide insertions or deletions within runs of five to seven As or Ts, although a deletion of 11 nucleotides also occurred, leading to frameshifts and premature stop codons. During 10 passage rounds, the attenuated VACV was replaced by the mutant viruses. At the end of the experiment, virtually all remaining viruses had one fixed mutation and one or more additional mutations. Although nucleotide substitutions in the transcription apparatus accounted for two low-frequency mutations, frameshifts in genes encoding protein components of the mature virion, namely, A26L, G6R, and A14.5L, achieved 74% to 98% fixation. The adaptive role of the mutations was confirmed by making recombinant VACV with A26L or G6R or both deleted, which increased virus replication levels and decreased particle/PFU ratios.IMPORTANCEGene inactivation is considered to be an important driver of orthopoxvirus evolution. Whereas cowpox virus contains intact orthologs of genes present in each orthopoxvirus species, numerous genes are inactivated in all other members of the genus. Inactivation of additional genes can occur upon extensive passaging of orthopoxviruses in cell culture leading to attenuationin vivo, a strategy for making vaccines. Whether inactivation of multiple viral genes enhances replication in the host cells or has a neutral effect is unknown in most cases. Using an experimental evolution protocol involving serial passages of an attenuated vaccinia virus, rapid acquisition of inactivating frameshift mutations occurred. After only 10 passage rounds, the starting attenuated vaccinia virus was displaced by viruses with one fixed mutation and one or more additional mutations. The high frequency of multiple inactivating mutations during experimental evolution simulates their acquisition during normal evolution and extensive virus passaging to make vaccine strains.
DOI：
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发表时间：
1977
期刊：
影响因子：
--
作者：
G. Avala
通讯作者：
G. Avala
中枢神经系统中的谷氨酰胺、谷氨酸和伽马胺编辑：L. Hertz、E. Kvamme、E.G. McGeer 和 A. Schousboe，纽约，1984 年。系列：神经病学和神经生物学，第 720 页。无花果和表格，66.00 英镑 ISBN 0-8451-2706-3。
DOI：
--
发表时间：
1986
期刊：
Clinical neurology and neurosurgery (Dutch-Flemish ed. Print)
影响因子：
--
作者：
A. Teelken
通讯作者：
A. Teelken