Rohll J B, Moon D H, Evans D J, Almond J W
Department of Microbiology, School of Animal and Microbial Sciences, University of Reading, Whiteknights, United Kingdom.
J Virol. 1995 Dec;69(12):7835-44. doi: 10.1128/JVI.69.12.7835-7844.1995.
The role of the 3' untranslated region (3'UTR) in the replication of enteroviruses has been studied with a series of mutants derived from either poliovirus type 3 (PV3) or a PV3 replicon containing the reporter gene chloramphenicol acetyltransferase. Replication was observed when the PV3 3'UTR was replaced with that of either coxsackie B4 virus, human rhinovirus 14 (HRV14), bovine enterovirus, or hepatitis A virus, despite the lack of sequence and secondary structure homology of the 3'UTRs of these viruses. The levels of replication observed for recombinants containing the 3'UTRs of hepatitis A virus and bovine enterovirus were lower than those for PV3 and the other recombinants. Extensive site-directed mutagenesis of the single stem-loop structure formed by the HRV14 3'UTR indicated the importance of (i) the loop sequence, (ii) the stability of the stem, and (iii) the location of the stem immediately upstream of the poly(A) tail. The role of a 4-bp motif at the base of the HRV14 stem, highly conserved among rhinoviruses, was examined by site-directed mutagenesis of individual base pairs. This analysis did not pinpoint a particular base pair as crucial for function. The requirement for immediate adjacent positioning of the open reading frame and the 3'UTR was examined by insertion of a 1.1-kb heterologous sequence. A replicon containing this insert replicated to about 30% of the level observed for the wild type. However, the corresponding virus consistently deleted most of the inserted fragment, suggesting that its presence was incompatible with a full replication cycle.
利用一系列源自3型脊髓灰质炎病毒（PV3）或含有报告基因氯霉素乙酰转移酶的PV3复制子的突变体，研究了3'非翻译区（3'UTR）在肠道病毒复制中的作用。当PV3的3'UTR被柯萨奇B4病毒、人鼻病毒14型（HRV14）、牛肠道病毒或甲型肝炎病毒的3'UTR取代时，尽管这些病毒的3'UTR缺乏序列和二级结构同源性，但仍观察到了复制现象。含有甲型肝炎病毒和牛肠道病毒3'UTR的重组体的复制水平低于PV3和其他重组体。对由HRV14 3'UTR形成的单茎环结构进行广泛的定点诱变表明：（i）环序列、（ii）茎的稳定性以及（iii）茎在多聚腺苷酸尾紧邻上游的位置很重要。通过对单个碱基对进行定点诱变，研究了HRV14茎基部高度保守于鼻病毒的4碱基基序的作用。该分析未确定某个特定碱基对对于功能至关重要。通过插入1.1 kb的异源序列，研究了开放阅读框和3'UTR紧邻定位的要求。含有该插入片段的复制子复制到野生型观察水平的约30%。然而，相应的病毒始终删除了大部分插入片段，表明其存在与完整的复制周期不相容。
