Evaluation of the role of heterogeneous nuclear ribonucleoprotein A1 as a host factor in murine coronavirus discontinuous transcription and genome replication.
Evaluation of the role of heterogeneous nuclear ribonucleoprotein A1 as a host factor in murine coronavirus discontinuous transcription and genome replication.
复制标题
评价异质核核糖核蛋白 A1 作为宿主因子在鼠冠状病毒不连续转录和基因组复制中的作用。
DOI：
10.1073/pnas.031424298
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发表时间：
2001
影响因子：
11.1
通讯作者：
Masters,PS
中科院分区：
文献类型：
--
作者：
Shen,X;Masters,PS
具有RNA基因组的病毒经常捕获和重定向宿主细胞成分，以辅助RNA依赖性RNA合成的特定机制。尼多病毒是正链RNA病毒的一个目，包括冠状病毒和动脉病毒，它们采用独特的不连续转录策略，产生一系列亚基因组mrna，将5 '先导体连接到基因组的远端部分。对于原型冠状病毒小鼠肝炎病毒（MHV），异质性核核糖核蛋白（hnRNP） A1已被证明能够在体外结合基因间序列的负链，这是一种在先导RNA中发现的顺式作用元件，位于基因组中每个下游ORF之前。因此，hnRNP A1被认为是MHV转录中的宿主因子。为了从遗传学上验证这一假设，我们最初构建了MHV突变体，将一个非常高亲和力的hnRNP A1结合位点插入MHV基因组中的基因间序列。插入的hnRNP A1结合位点不能在功能上取代或增强基因间序列的转录。这一发现使我们通过分析不表达该蛋白的小鼠细胞系中MHV的复制和RNA合成来更直接地测试hnRNP A1的作用。细胞中hnRNP A1的缺失对感染性病毒的产生、基因组RNA的合成或亚基因组mrna的数量和质量没有可检测到的影响。这些结果强烈提示hnRNP A1不是MHV不连续转录或基因组复制所需的宿主因子。
Viruses with RNA genomes often capture and redirect host cell components to assist in mechanisms particular to RNA-dependent RNA synthesis. The nidoviruses are an order of positive-stranded RNA viruses, comprising coronaviruses and arteriviruses, that employ a unique strategy of discontinuous transcription, producing a series of subgenomic mRNAs linking a 5′ leader to distal portions of the genome. For the prototype coronavirus mouse hepatitis virus (MHV), heterogeneous nuclear ribonucleoprotein (hnRNP) A1 has been shown to be able to bindin vitroto the negative strand of the intergenic sequence, a cis-acting element found in the leader RNA and preceding each downstream ORF in the genome. hnRNP A1 thus has been proposed as a host factor in MHV transcription. To test this hypothesis genetically, we initially constructed MHV mutants with a very high-affinity hnRNP A1 binding site inserted in place of, or adjacent to, an intergenic sequence in the MHV genome. This inserted hnRNP A1 binding site was not able to functionally replace, or enhance transcription from, the intergenic sequence. This finding led us to test more directly the role of hnRNP A1 by analysis of MHV replication and RNA synthesis in a murine cell line that does not express this protein. The cellular absence of hnRNP A1 had no detectable effect on the production of infectious virus, the synthesis of genomic RNA, or the quantity or quality of subgenomic mRNAs. These results strongly suggest that hnRNP A1 is not a required host factor for MHV discontinuous transcription or genome replication.