Boyce Thompson Institute at Cornell University, Ithaca, New York 14853, USA.
Virology. 2011 Jul 20;416(1-2):54-64. doi: 10.1016/j.virol.2011.04.019. Epub 2011 May 23.
To examine the role of the AcMNPV lef-5 gene in the context of the infection cycle, we generated an AcMNPV lef-5 knockout virus (vAc(lef5ko)) and a complementing cell line that supports viral replication. We examined AcMNPV DNA replication, early and late gene expression, and production of infectious viral progeny in the absence of lef-5. While early gene expression and DNA replication were not reduced by the lef-5 knockout, expression of a late reporter was disrupted and representative late transcripts were dramatically reduced. Progeny virus production was not detected after transfection of Sf9 cells with the lef-5 knockout bacmid, but was rescued by insertion of an egfp- or myc-tagged lef-5 gene into the vAc(lef5ko) genome. An egfp-tagged lef-5 gene from SeMNPV was used to generate a stable Sf9 cell line that supported replication of the vAc(lef5ko) virus. The LEF-5 protein was also found to co-localize with IE-1 in infected cell nuclei.
为了研究 AcMNPV lef-5 基因在感染周期中的作用，我们构建了 AcMNPV lef-5 基因敲除病毒（vAc(lef5ko)）和支持病毒复制的互补细胞系。我们检测了 AcMNPV DNA 复制、早期和晚期基因表达以及在没有 lef-5 的情况下产生感染性病毒后代的情况。虽然 lef-5 基因敲除不会降低早期基因表达和 DNA 复制，但晚期报告基因的表达受到干扰，代表性的晚期转录本显著减少。用 lef-5 基因敲除杆状病毒转染 Sf9 细胞后，无法检测到后代病毒的产生，但将 egfp-或 myc 标记的 lef-5 基因插入 vAc(lef5ko)基因组中可挽救后代病毒的产生。我们还使用 SeMNPV 的 egfp 标记 lef-5 基因构建了一个稳定的 Sf9 细胞系，该细胞系支持 vAc(lef5ko)病毒的复制。在感染细胞的核内还发现 LEF-5 蛋白与 IE-1 共定位。
