Reverse genetics of negative-strand RNA viruses: closing the circle.
Reverse genetics of negative-strand RNA viruses: closing the circle.
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DOI：
10.1073/pnas.96.16.8804
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发表时间：
1999-08
影响因子：
11.1
通讯作者：
Andrew Pekosz;Biao He;Robert A. Lamb
中科院分区：
文献类型：
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作者：
Andrew Pekosz;Biao He;Robert A. Lamb
对病毒及其与宿主细胞和生物体相互作用的研究极大地受益于将特定突变设计到病毒基因组中的能力，这种技术被称为反向遗传学。DNA病毒的基因组操作是最先进行的，通过用编码病毒基因组的质粒转染细胞（1）或通过携带病毒序列的质粒与病毒基因组的异源重组（2-4）。正链RNA病毒基因组操作紧随其后，部分原因是病毒基因组也是mRNA正义的。简单地将含有脊髓灰质炎病毒基因组的质粒或从质粒转录的RNA转移到易感细胞中导致感染性脊髓灰质炎病毒的回收（5，6）。负链RNA病毒包括许多人类和动物病原体，例如流感A、B和C病毒、汉坦病毒、拉沙病毒、狂犬病病毒、埃博拉病毒、马尔堡病毒、麻疹病毒、犬瘟热病毒、牛瘟病毒、呼吸道合胞病毒、腮腺炎病毒、人副流感病毒1-4型和尼帕病毒（最近在马来西亚出现，导致猪和人的呼吸窘迫和脑炎）。然而，负链RNA病毒的基因组不太适合人工操作，原因如下：（i）基因组RNA的复制和包装需要精确的5′和3′末端;（ii）病毒RNA聚合酶对于转录mRNA和互补的正义反基因组模板RNA都是必需的;和（iii）基因组和反基因组RNA都以病毒核糖核蛋白（RNP）复合物的形式存在（参考文献7）。流感病毒、布尼亚病毒和沙粒病毒的分段基因组允许通过分离重组病毒进行一些遗传操作，但是分段负链RNA病毒的完整基因组的操作进展缓慢，受到基因组被分段的事实的阻碍。
The study of viruses and their interactions with host cells and organisms has benefited greatly from the ability to engineer specific mutations into viral genomes, a technique known as reverse genetics. Genome manipulations of DNA viruses, either by transfecting cells with plasmids encoding the viral genome (1) or by heterologous recombination of plasmids bearing viral sequences with the virus genome (2–4), were the first to be performed. Positive-strand RNA virus genome manipulation followed quickly, partly because the viral genome is also mRNA sense. Simply transfecting plasmids, or RNA transcribed from plasmids, containing the poliovirus genome into susceptible cells resulted in the recovery of infectious poliovirus (5, 6). The negative-strand RNA viruses include a number of human and animal pathogens such as influenza A, B, and C viruses, hantaviruses, Lassa virus, rabies virus, Ebola virus, Marburg virus, measles virus, canine distemper virus, rinderpest virus, respiratory syncytial virus, mumps virus, human parainfluenza virus types 1–4, and Nipah virus (which recently emerged in Malaysia, causing respiratory distress and encephalitis in pigs and humans). However, the genomes of the negative-strand RNA viruses have been less amenable to artificial manipulation for several reasons: (i) precise 5′ and 3′ ends are required for replication and packaging of the genomic RNA; (ii) the viral RNA polymerase is essential for transcribing both mRNA and complementary, positive-sense antigenome template RNA; and (iii) both genomic and antigenomic RNAs exist as viral ribonucleoprotein (RNP) complexes (reviewed in ref. 7). The segmented genomes of influenza viruses, bunyaviruses, and arenaviruses allowed some genetic manipulation through the isolation of reassortant viruses, but manipulation of the complete genome of segmented negative-strand RNA viruses has progressed slowly, hampered by the very fact that the genome is segmented.