THE HR5 TRANSCRIPTIONAL ENHANCER STIMULATES EARLY EXPRESSION FROM THE AUTOGRAPHA-CALIFORNIA NUCLEAR POLYHEDROSIS-VIRUS GENOME BUT IS NOT REQUIRED FOR VIRUS-REPLICATION
THE HR5 TRANSCRIPTIONAL ENHANCER STIMULATES EARLY EXPRESSION FROM THE AUTOGRAPHA-CALIFORNIA NUCLEAR POLYHEDROSIS-VIRUS GENOME BUT IS NOT REQUIRED FOR VIRUS-REPLICATION
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DOI：
10.1128/jvi.67.10.5776-5785.1993
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发表时间：
1993-10-01
影响因子：
5.4
通讯作者：
FRIESEN, PD
中科院分区：
文献类型：
--
作者：
RODEMS, SM;FRIESEN, PD
苜蓿银纹夜蛾核型多角体病毒（Autographa californica nuclear polyhedrosis virus，AcMNPV）基因组中分布着5个同源区（hr 1 ~ hr 5）。质粒转染的分析表明，hrs的功能作为转录增强子和病毒DNA复制的可能起源。通过构建一系列重组病毒来检测这些重复元件在调节AcMNPV基因组表达中的作用，所述重组病毒测试hr 5对相邻p35基因（p35）表达的影响。当嵌入病毒基因组中时，hr 5以位置和方向独立的方式刺激早期p35启动子的转录。此外，hr 5和p35的上游激活区在功能上是可互换的。一个28 bp的不完美回文，重复6次内hr 5，是最小的序列所需的p35启动子激活。hr 5也刺激另一个早期AcMNPV启动子，但不是一个晚期启动子或宿主衍生的启动子，这表明增强是启动子特异性感染期间。为了研究其在AcMNPV复制过程中的作用，将hr 5从病毒基因组内的正常位置删除。由此产生的hr 5突变体没有表现出明显的复制缺陷，作为判断生产的出芽病毒和非常晚的基因表达水平，即使稳定状态的p35 RNA水平降低。这些结果首次表明hr 5在病毒基因组中起转录增强子的作用。然而，该元件不是培养细胞中AcMNPV复制所必需的。因此，DNA复制的五个可能起点之一的丢失对病毒生长无害。由于p26从hr 5缺失突变体中去除，因此该基因对于病毒复制也是非必需的。
Autographa californica nuclear polyhedrosis virus (AcMNPV) contains five homologous regions (hr1 through hr5) interspersed throughout its genome. Analysis of plasmid transfections indicates that the hrs function as transcriptional enhancers and possible origins of viral DNA replication. The role of these repetitive elements in regulating expression from the AcMNPV genome was examined by constructing a series of recombinant viruses that tested the effect of hr5 on expression of the adjacent p35 gene (p35). When embedded within the viral genome, hr5 stimulated transcription from the early p35 promoter in a position- and orientation-independent manner. Moreover, hr5 and the upstream activating region of p35 were functionally interchangeable. A 28-bp imperfect palindrome, repeated six times within hr5, was the minimal sequence required for p35 promoter activation. hr5 also stimulated another early AcMNPV promoter but not a late promoter or a host-derived promoter, suggesting that enhancement is promoter specific during infection. To investigate its role during AcMNPV replication, hr5 was deleted from its normal position within the viral genome. The resulting hr5 mutants exhibited no apparent defects in replication, as judged by production of budded virus and levels of very late gene expression, even though steady-state levels of p35 RNA were reduced. These results indicated for the first time that hr5 functions as a transcriptional enhancer within the viral genome. However, the element is not required for AcMNPV replication in cultured cells. Thus, loss of one of five possible origins of DNA replication is not deleterious to viral growth. Since p26 was removed from the hr5 deletion mutants, this gene is also nonessential for viral replication.