RNA replication from the simian virus 5 antigenomic promoter requires three sequence-dependent elements separated by sequence-independent spacer regions.
RNA replication from the simian virus 5 antigenomic promoter requires three sequence-dependent elements separated by sequence-independent spacer regions.
复制标题
猿猴病毒 5 反基因组启动子的 RNA 复制需要三个由序列无关的间隔区分隔的序列依赖性元件。
DOI：
10.1128/jvi.75.8.3993-3998.2001
复制
发表时间：
2001
影响因子：
5.4
通讯作者：
Parks,GD
中科院分区：
文献类型：
--
作者：
Keller,MA;Murphy,SK;Parks,GD
我们之前已经证明，对于副粘病毒猿猴病毒5（SV5），RNA复制的功能性启动子需要两个不连续元件之间的适当间隔：3′端的19个碱基片段（保守区I [CRI]）和L蛋白基因编码区内包含的18个碱基的内部区域（CRII）。在这里描述的工作中，我们使用了反向遗传学系统来确定CRI和CRII之间的53个碱基片段是否包含最佳复制所需的额外序列特异性信号，或者该片段是否仅作为序列独立的间隔区发挥作用。构建了一系列复制回缺陷干扰微型基因组类似物，以含有非病毒序列取代反基因组启动子的碱基21至72，并通过北方印迹分析测量RNA复制的相对水平。我们的突变分析结果表明，除了CRI和CRII之外，SV5反基因组启动子的最佳复制需要位于距离RNA 3′端51至66个碱基的第三个序列依赖元件。微型基因组RNA复制不受影响的变化，无论是在该元件的位置，CRI和CRII或预测的NP的六聚体阶段。因此，从SV5反基因组启动子的最佳RNA复制需要三个序列依赖性元件，CRI，CRII和碱基51至66。
We have previously shown for the paramyxovirus simian virus 5 (SV5) that a functional promoter for RNA replication requires proper spacing between two discontinuous elements: a 19-base segment at the 3′ terminus (conserved region I [CRI]) and an 18-base internal region (CRII) that is contained within the coding region of the L protein gene. In the work described here, we have used a reverse-genetics system to determine if the 53-base segment between CRI and CRII contains additional sequence-specific signals required for optimal replication or if this segment functions solely as a sequence-independent spacer region. A series of copyback defective interfering minigenome analogs were constructed to contain substitutions of nonviral sequences in place of bases 21 to 72 of the antigenomic promoter, and the relative level of RNA replication was measured by Northern blot analysis. The results from our mutational analysis indicate that in addition to CRI and CRII, optimal replication from the SV5 antigenomic promoter requires a third sequence-dependent element located 51 to 66 bases from the 3′ end of the RNA. Minigenome RNA replication was not affected by changes in the either the position of this element in relation to CRI and CRII or the predicted hexamer phase of NP encapsidation. Thus, optimal RNA replication from the SV5 antigenomic promoter requires three sequence-dependent elements, CRI, CRII and bases 51 to 66.