Positive- and negative-acting signals combine to determine differential RNA replication from the paramyxovirus simian virus 5 genomic and antigenomic promoters.
Positive- and negative-acting signals combine to determine differential RNA replication from the paramyxovirus simian virus 5 genomic and antigenomic promoters.
复制标题
正作用和负作用信号结合起来确定副粘病毒猿病毒5基因组和反基因组启动子的RNA复制差异。
DOI：
10.1016/s0042-6822(02)00071-5
复制
发表时间：
2003
期刊：
影响因子：
3.7
通讯作者：
Parks,GriffithD
中科院分区：
文献类型：
--
作者：
Keller,MichaelA;Parks,GriffithD
在基因组和反基因组RNA的3′端发现的顺式作用信号是决定副粘病毒RNA从每个启动子复制水平的主要因素。使用一个微型基因组系统，重建SV 5 RNA合成从cDNA衍生的组件，我们在这里显示，副粘病毒SV 5的基因组启动子（GP）指导RNA复制比从反基因组启动子（AGP）看到的低14倍。本研究的目的是鉴定负责SV 5 GP和AGP RNA复制差异水平的顺式作用信号。我们先前已经表明，SV 5 AGP包含三个序列依赖性元件（CRI，CRII和区域III），它们被序列独立的间隔区分开。构建了含有嵌合启动子的微型基因组，以检验将离散的顺式作用AGP元件转移到GP可以赋予GP更高的复制特性的假设。含有AGP CRI、CRII或区域III元件单独取代相应GP序列的微型基因组未显示出RNA复制水平的增强。然而，将AGP 3′末端CRI和区域III元件转移到GP的相应位点导致了一个微型基因组，其复制水平为AGP的约40%。通过还包括位于CRI和区域III之间的插入AGP区段（碱基20-50），来自GP的这种增强的RNA复制进一步增加至AGP水平。重要的是，转移非病毒序列代替GP碱基20-50也增加RNA复制到接近AGP的水平，但仅在AGP CRI和区域III取代的情况下。这些数据表明，来自SV 5 GP和AGP的RNA复制水平的差异是由于AGP（CRI和区域III）中的正作用信号和GP（碱基20-50）中的负作用信号的组合。SV 5启动子元件在确定RNA复制水平的可能功能提出。
The cis-acting signals found at the 3′ ends of the genomic and antigenomic RNAs are a major factor determining the level of paramyxovirus RNA replication from each promoter. Using a minigenome system that reconstitutes SV5 RNA synthesis from cDNA-derived components, we show here that the genomic promoter (GP) for the paramyxovirus SV5 directs RNA replication ∼14-fold lower than that seen from the antigenomic promoter (AGP). The goal of this study was to identify cis-acting signals responsible for differential levels of RNA replication from the SV5 GP and AGP. We have previously shown that the SV5 AGP contains three sequence-dependent elements (CRI, CRII, and Region III) that are separated by sequence-independent spacer regions. Minigenomes containing chimeric promoters were constructed to test the hypothesis that transfer of discrete cis-acting AGP elements to the GP could confer higher replication properties to the GP. Minigenomes containing a substitution of the AGP CRI, CRII, or Region III elements alone in place of the corresponding GP sequences did not show enhanced levels of RNA replication. However, transfer of both the AGP 3′ terminal CRI and Region III elements into the corresponding sites of the GP led to a minigenome which replicated to ∼40% of the levels seen with the AGP. This enhanced RNA replication from the GP was further increased up to AGP levels by also including the intervening AGP segment (bases 20–50) located between CRI and Region III. Importantly, transfer of nonviral sequences in place of GP bases 20–50 also increased RNA replication to levels approaching that of the AGP, but only in the context of the AGP CRI and Region III substitutions. These data indicate that differential levels of RNA replication from the SV5 GP and AGP are due to a combination of positive-acting signals in the AGP (CRI and Region III) and a negative-acting signal in the GP (bases 20–50). Possible functions for the SV5 promoter elements in determining RNA replication levels are proposed.