The Role of 5' ICR Sequences in Viral (+) RNA Replication

人类、牲畜和农作物的大多数病毒性疾病都是由具有RNA基因组的病毒引起的。尽管具有明显的重要性，但对RNA病毒复制的分子过程所知甚少。重组DNA技术已经提供了解决这个问题的方法，其中之一就是设计病毒rna的cDNA拷贝的能力。从野生型病毒序列的cdna在体外转录RNA，在一些情况下，产生感染性RNA。cDNA水平上的突变相对容易，这种改变的克隆的RNA转录物可以作为RNA水平上的特定突变体。最近在雀麦花叶病毒（BMV）和其他病毒的基因组rna的5'末端发现了与tRNA基因的内部启动子序列惊人相似的内部控制区（ICRs），也称为A盒和B盒。5'端位置，它们与最近表征的亚基因组启动子核心序列的部分同源性，以及BMV基因组RNA3的缺失分析提供了强有力的间接证据，证明这些序列是（+）链复制的启动子。Pi使用野生型和突变转录本结合独特的体外复制程序来定义基因组（-）链合成和亚基因组（+）链合成的启动子。然而，（+）链基因组RNA合成的启动子尚未明确。在体内，病毒RNA的合成通常是高度不对称的，（+）链RNA的合成比（-）链多出许多倍。n tRNA基因在真核生物中的转录是一个核过程，依赖于聚合酶III和相关转录因子。与trna启动子的明显同源性表明宿主因子参与了病毒（+）链的合成，可能是pol III转录因子。实验是为了确定在BMV的ICRs内和周围的哪些元素在复制中是必不可少的和起作用的。此外，ICRs在复制不对称中的作用，以及在确定基因组rna合成的特定摩尔比方面的作用。鉴定参与复制的宿主因子是至关重要的，因为它们是宿主-病原体关系的核心。

Most viral diseases, of man, his livestock, and crops are caused by viruses having RNA genomes. Despite the obvious importance, remarkably little is known about the molecular processes of RNA viral replication. Recombinant DNA techniques have provided procedures for addressing the problem, one of these being the ability to engineer cDNA copies of viral RNAs. Transcription of RNA in vitro from cDNAs of wild-type viral sequences has, in several cases, yielded infectious RNA. Mutagenesis at the cDNA level is relatively facile, and RNA transcripts of such altered clones can serve as specific mutants at the RNA level. Sequences with striking resemblance to the internal promoters of tRNA genes, internal control regions (ICRs), also termed A and B boxes, have recently been observed at the 5' termini of the genomic RNAs of brome mosaic virus (BMV)and other viruses. The 5' terminal location, their partial homology to the recently characterized core sequence f the subgenomic promoter, and the deletion analysis of BMV genomic RNA3 provide strong circumstantial evidence that these sequenced function as promoters of (+) strand replication. The Pi has used wild type and mutant transcripts in combination with unique in vitro replication procedures to define the promoters of genomic (-) strand synthesis and subgenomic (+) strand synthesis. However, the promoters of (+) strand genomic RNA synthesis are as yet undefined. In vivo viral RNA synthesis is usually highly asymmetric with many times more (+) strand RNA being synthesized than (-) strand.n tRNA gene transcription in eukaryotes, dependent on polymerase III and associated transcription factors, is a nuclear process. The apparent homology to promoters of tRNAs suggests involvement of host factors in viral (+) strand synthesis, possibly pol III transcription factors are proposed. Experiments are to define which elements within and surrounding the ICRs of BMV are essential and functional in replication. Furthermore, the role of the ICRs in the asymmetry of replication, and in determining the specific molar ratios of genomic RNAs synthesized is addressed. Identification of the host factor(s) involved in replication is essential in that they are the core of the host-pathogen relationship.