Weiss B G, Schlesinger S
Department of Molecular Microbiology, Washington University School of Medicine, St. Louis, Missouri 63110-1093.
J Virol. 1991 Aug;65(8):4017-25. doi: 10.1128/JVI.65.8.4017-4025.1991.
The genome (49S RNA) of Sindbis virus is a positive-strand RNA of 11.7 kb that consists of two domains. The 5' two-thirds of the RNA codes for the proteins required for replication and transcription of the RNA. The 3' one-third codes for the structural proteins. The latter are translated from a 26S subgenomic RNA identical in sequence to the 3' one-third of the genome. The 26S RNA is transcribed by initiation from an internal promoter that spans the junction between the nonstructural and structural genes. We have used Sindbis virus RNAs transcribed from cloned cDNAs to demonstrate recombination between Sindbis virus RNAs in cultured cells. Several different combinations of deleted or mutationally altered RNAs gave rise to infectious recombinants. In 7 of 10 different crosses, the infectious recombinant RNAs were larger than wild-type 49S RNA. We sequenced the recombinant RNAs in the region spanning the junction between the nonstructural and structural protein genes from five different crosses. In three of the crosses, this is the only region within which recombination could have taken place to produce an infectious 49S RNA. Recombination also occurred in this region in the other two crosses. The recombinant RNAs were distinct from wild-type RNA and from each other. All contained sequence insertions derived from the parental RNAs. One contained a deletion and a rearrangement, and one also contained a stretch of 11 nucleotides not found in the Sindbis virus genome. When each of the parental RNAs contained a functional subgenomic RNA promoter, both promoters were present and functional in the recombinant RNA. Those recombinants with large sequence insertions showed evidence of evolution toward the wild-type single-junction RNA.
辛德毕斯病毒的基因组（49S RNA）是一条11.7 kb的正链RNA，由两个结构域组成。RNA 5'端的三分之二编码RNA复制和转录所需的蛋白质。3'端的三分之一编码结构蛋白。后者由一个26S亚基因组RNA翻译而来，该RNA的序列与基因组3'端的三分之一相同。26S RNA通过跨越非结构基因和结构基因之间连接处的内部启动子起始转录。我们使用从克隆的cDNA转录而来的辛德毕斯病毒RNA，来证明培养细胞中辛德毕斯病毒RNA之间的重组。几种不同的缺失或突变改变的RNA组合产生了感染性重组体。在10个不同的杂交组合中，有7个组合产生的感染性重组RNA比野生型49S RNA大。我们对来自五个不同杂交组合的、跨越非结构蛋白基因和结构蛋白基因之间连接处的区域的重组RNA进行了测序。在其中三个杂交组合中，这是唯一可能发生重组以产生感染性49S RNA的区域。另外两个杂交组合在这个区域也发生了重组。重组RNA与野生型RNA不同，彼此之间也不同。所有重组RNA都包含来自亲本RNA的序列插入。一个包含一个缺失和一个重排，还有一个包含一段在辛德毕斯病毒基因组中未发现的11个核苷酸的片段。当每个亲本RNA都包含一个功能性亚基因组RNA启动子时，两个启动子在重组RNA中都存在且具有功能。那些具有大序列插入的重组体显示出向野生型单连接RNA进化的证据。
