Qiu W, Liu J J, Carstens E B
Department of Microbiology and Immunology, Queen's University, Kingston, Ontario, Canada.
Virology. 1996 Mar 15;217(2):564-72. doi: 10.1006/viro.1996.0151.
To investigate the mechanisms regulating baculovirus virulence and host range we have begun to study Choristoneura fumiferana nuclear polyhedrosis virus (CfMNPV) and its gene expression in permissive and nonpermissive cells. We have identified and mapped three genes on the CfMNPV genome. The polyhedrin gene is located from 0.0 to 2.0 m.u. and two other genes, dnapol and p143, both of which are essential for baculovirus DNA replication, are located from 35.3 to 40.9 m.u. and 55.5 to 63.4 m.u., respectively. To gain insight into the expression of CfMNPV genes in permissive C. fumiferana and nonpermissive Spodoptera frugiperda cells, we constructed three expression plasmids in which the promoter region of the dnapol, the p143, and polyhedrin genes were placed in front of a chloramphenicol acetyltransferase reporter gene. All three CfMNPV promoters were active in nonpermissive cells in the presence of Autographa californica nuclear polyhedrosis virus (AcMNPV) DNA, but no activity was detected in permissive cells either in the presence of CfMNPV DNA or AcMNPV DNA. This lack of promoter activity was not due to failure of viral or plasmid DNA to enter the cell nucleus. It was possible that the reporter plasmids were inefficient templates for transcriptional transactivation so we developed a CfMNPV transfer vector and generated a recombinant virus in which the polyhedrin promoter driving CAT gene cassette was integrated into the CfMNPV genome. In this case, the CfMNPV polyhedrin promoter was highly active in the permissive cells.
为了研究调节杆状病毒毒力和宿主范围的机制，我们已开始研究云杉卷叶蛾核型多角体病毒（CfMNPV）及其在允许细胞和非允许细胞中的基因表达。我们已在CfMNPV基因组上鉴定并定位了三个基因。多角体蛋白基因位于0.0至2.0个图距单位处，另外两个对杆状病毒DNA复制至关重要的基因，即dnapol和p143，分别位于35.3至40.9个图距单位处和55.5至63.4个图距单位处。为了深入了解CfMNPV基因在允许的云杉卷叶蛾细胞和非允许的草地贪夜蛾细胞中的表达情况，我们构建了三个表达质粒，其中dnapol、p143和多角体蛋白基因的启动子区域位于氯霉素乙酰转移酶报告基因之前。在苜蓿银纹夜蛾核型多角体病毒（AcMNPV）DNA存在的情况下，所有三个CfMNPV启动子在非允许细胞中均有活性，但在存在CfMNPV DNA或AcMNPV DNA的允许细胞中均未检测到活性。这种启动子活性的缺乏并非由于病毒或质粒DNA未能进入细胞核。有可能报告质粒是转录反式激活的低效模板，因此我们开发了一种CfMNPV转移载体，并产生了一种重组病毒，其中驱动CAT基因盒的多角体蛋白启动子被整合到CfMNPV基因组中。在这种情况下，CfMNPV多角体蛋白启动子在允许细胞中具有高度活性。
