Characterization of Bunyamwera virus defective interfering particles.
Characterization of Bunyamwera virus defective interfering particles.
复制标题
Bunyamwera 病毒缺陷干扰颗粒的表征。
DOI：
10.1099/0022-1317-73-2-389
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发表时间：
1992
影响因子：
3.8
通讯作者：
R. Elliott
中科院分区：
文献类型：
--
作者：
A. H. Patel;R. Elliott
为了分离出条件致死的布尼安维拉病毒琥珀无义突变体，在BHK和小鼠L细胞上发现了5个能产生小斑块的变异株。Northern blotting分析表明，这些变异体合成了来自L核糖核酸片段的缺陷(亚基因组)RNA。未检测到亚基因组M和S片段RNA。有缺陷的L RNA被包装成病毒颗粒，五种制剂中有四种对标准病毒的增殖造成了干扰。当含有缺陷的制剂与标准病毒混合并在双重感染的细胞中生长时，观察到标准病毒的滴度最多减少了400倍。因此，这些制剂很可能含有缺陷干扰(DI)颗粒。在DI病毒感染的细胞中合成了新的DI特异性多肽。这些新蛋白可以被针对L蛋白N端和/或C端的抗血清沉淀。对3种不同缺陷病毒的主要DIRNA克隆的核苷酸序列分析表明，每例DIRNA都发生了L片段的单一内部缺失，但保留了5‘端和3’端的序列。L基因片段的缺失程度在72%到77%之间。我们的结果提示，这些DI颗粒可能是在试图从小鼠L细胞上分离本安韦拉病毒琥珀突变体时产生的，因为在野生型病毒感染后，来自L和M节段的缺陷/亚基因组RNA很容易在小鼠L细胞中产生，而在BHK细胞中则不产生。
In an attempt to isolate conditional lethal amber nonsense mutants of Bunyamwera virus, five variants were found which produced small plaques on BHK and mouse L cells. Characterization of these variants by Northern blotting showed that they synthesized defective (subgenomic) RNAs derived from the L RNA segment. No subgenomic M or S segment RNAs were detected. The defective L RNAs were shown to be packaged into virus particles, and four of five preparations caused interference with the multiplication of standard virus. When defective-containing preparations were mixed with standard virus and grown in doubly infected cells a reduction in titre of standard virus of up to 400-fold was observed. Hence these preparations most probably contained defective interfering (DI) particles. Novel DI-specific polypeptides were synthesized in DI virus-infected cells. These novel proteins could be precipitated by antisera raised against either the N or C terminus, or both, of the L protein. Nucleotide sequence analysis of cloned cDNA to prominent DI RNAs in three different defective virus preparations revealed that the DI RNA in each case had suffered a single internal deletion of the L segment while retaining the 5'- and 3'-terminal sequences. The extent of the deletion ranged between 72% and 77% of the L RNA segment. Our results suggest that these DI particles may have arisen during the attempted isolation of Bunyamwera virus amber mutants on mouse L cells, since defective/subgenomic RNAs derived from the L and M segments were readily generated in mouse L cells but not in BHK cells, following infection with wild-type virus.