Huang Jianhe, Levin David B
Department of Biology, University of Victoria, PO Box 3020, Victoria, British Columbia, Canada V8W 3N51.
J Gen Virol. 1999 Aug;80 ( Pt 8):2263-2274. doi: 10.1099/0022-1317-80-8-2263.
A putative non-hr origin of DNA replication was identified in the Spodoptera littoralis multinucleocapsid nucleopolyhedrovirus (SpliNPV) genome by transient replication assays. The putative SpliNPV ori was mapped to the PstI-J fragment between 75.1-77.9 map units in the SpliNPV genome. While the DNA sequence of the putative SpliNPV ori aligned with regions within the non-hr oris of Autographa californica, Orgyia pseudotsugata and Spodoptera exigua multinucleocapsid nucleopolyhedroviruses, it has limited DNA sequence identity with these elements. The sequence of the putative SpliNPV non-hr ori fragment contains a unique distribution of imperfect palindromes, multiple direct repeats and putative transcription factor-binding sites. Transient expression assays indicated that the putative SpliNPV ori fragment repressed SpliNPV lef-3 promoter-mediated luciferase reporter gene expression. However, the putative SpliNPV ori fragment itself was capable of directing luciferase expression in the absence of a recognizable baculovirus promoter element in an orientation-independent fashion, suggesting that DNA sequence motifs within its sequence can activate transcription. Gel mobility shift analyses confirmed that proteins within nuclear extracts from both uninfected and virus-infected cells bound with specificity to the putative SpliNPV ori fragment.
通过瞬时复制试验，在斜纹夜蛾多核衣壳核多角体病毒（SpliNPV）基因组中鉴定出一个假定的非同源重复序列（non-hr）DNA复制起始位点。假定的SpliNPV ori被定位到SpliNPV基因组中75.1 - 77.9图谱单位之间的PstI-J片段。虽然假定的SpliNPV ori的DNA序列与苜蓿银纹夜蛾、云杉芽卷蛾和甜菜夜蛾多核衣壳核多角体病毒的非同源重复序列区域对齐，但其与这些元件的DNA序列同一性有限。假定的SpliNPV非同源重复序列ori片段的序列包含不完全回文、多个直接重复和假定的转录因子结合位点的独特分布。瞬时表达试验表明，假定的SpliNPV ori片段抑制了SpliNPV lef-3启动子介导的荧光素酶报告基因表达。然而，假定的SpliNPV ori片段本身能够在没有可识别的杆状病毒启动子元件的情况下以方向独立的方式指导荧光素酶表达，这表明其序列中的DNA序列基序可以激活转录。凝胶迁移率变动分析证实，未感染和病毒感染细胞的核提取物中的蛋白质都能特异性结合假定 的SpliNPV ori片段。
