Nalçacioğlu Remziye, Ince Ikbal Agah, Vlak Just M, Demirbağ Zihni, van Oers Monique M
Department of Biology, Faculty of Arts and Sciences, Karadeniz Technical University, 61080, Trabzon, Turkey.
Laboratory of Virology, Wageningen University, Binnenhaven 11, Wageningen, 6709 PD, The Netherlands.
J Gen Virol. 2007 Sep;88(Pt 9):2488-2494. doi: 10.1099/vir.0.82947-0.
The delayed-early DNA polymerase promoter of Chilo iridescent virus (CIV), officially known as Invertebrate iridescent virus, was fine mapped by constructing a series of increasing deletions and by introducing point mutations. The effects of these mutations were examined in a luciferase reporter gene system using Bombyx mori cells transfected with promoter constructs and infected with CIV. When the size of the upstream element was reduced from position -19 to -15, relative to the transcriptional start site, the luciferase activity was reduced to almost zero. Point mutations showed that each of the 5 nt (AAAAT) located between -19 and -15 were equally essential for promoter activity. Mutations at individual bases around the transcription initiation site showed that the promoter extended until position -2 upstream of the transcription start site. South-Western analysis showed that a protein of approximately 100 kDa interacted with the -19 nt promoter fragment in CIV-infected cells. This binding did not occur with a point mutant that lacked promoter activity. The AAAAT motif was also found in the DNA polymerase promoter region of other iridoviruses and in other putative CIV delayed-early genes.
稻纵卷叶螟虹彩病毒（CIV，正式名称为无脊椎动物虹彩病毒）的延迟早期DNA聚合酶启动子，通过构建一系列递增缺失片段和引入点突变进行了精细定位。在使用转染了启动子构建体并感染了CIV的家蚕细胞的荧光素酶报告基因系统中，检测了这些突变的影响。当相对于转录起始位点，上游元件的大小从-19位减少到-15位时，荧光素酶活性几乎降至零。点突变表明，位于-19和-15之间的5个核苷酸（AAAAT）中的每一个对于启动子活性都同样重要。转录起始位点周围单个碱基的突变表明，启动子延伸到转录起始位点上游-2位。蛋白质印迹分析表明，在CIV感染的细胞中，一种约100 kDa的蛋白质与-19 nt启动子片段相互作用。这种结合在缺乏启动子活性的点突变体中未发生。在其他虹彩病毒的DNA聚合酶启动子区域以及其他假定的CIV延迟早期基因中也发现了AAAAT基序。
