Bombyx mori nucleopolyhedrovirus ORF40 is essential for budded virus production and occlusion-derived virus envelopment
Bombyx mori nucleopolyhedrovirus ORF40 is essential for budded virus production and occlusion-derived virus envelopment
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家蚕核多角体病毒 ORF40 对于出芽病毒的产生和包膜病毒的包膜至关重要
DOI：
10.1099/jgv.0.001066
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发表时间：
2018
影响因子：
3.8
通讯作者：
Wu Xiaofeng
中科院分区：
文献类型：
--
作者：
Shen Yunwang;Feng Min;Wu Xiaofeng
家蚕核型多角体病毒（BmNPV）的ORF 40（bm40）编码苜蓿银纹夜蛾核型多角体病毒（AcMNPV）AC51的同源基因，是已测序的α杆状病毒中高度保守的基因。为了研究bm40基因在杆状病毒感染过程中的作用，通过在大肠杆菌中同源重组的方法构建了bm40基因敲除的BmNPV杆状病毒杆状病毒质粒。Western blotting分析表明，bm 40是病毒感染过程中的晚期基因。与野生型和修复病毒相比，敲除病毒表现出单细胞感染表型。滴定分析证实，由于bm40缺失，没有产生感染性出芽病毒（BV），同时对病毒DNA复制没有影响。电子显微镜显示，Bm40是所需的核衣壳出口从细胞核到细胞质，核衣壳形成闭塞衍生的病毒（ODV），并随后嵌入到多面体的ODV。共聚焦显微镜观察发现，Bm40在感染后48 h开始主要定位于细胞核内，感染后期主要聚集在核膜和多角体上。综上所述，这些结果表明，Bm40在BmNPV感染周期中BV产生和ODV表达中起重要作用。
ORF40 (bm40) of the Bombyx mori nucleopolyhedrovirus (BmNPV) encodes a homologue of Autographa califomica multiple nucleopolyhedrovirus (AcMNPV) AC51 and is a highly conserved gene in sequenced alphabaculoviruses. To investigate the role of bm40 in the baculovirus infection cycle, a bm40 knockout BmNPV bacmid was constructed via homologous recombination in Escherichia coil. Western blotting analysis revealed that bm40 is a late gene during virus infection. Compared with wild-type and repair viruses, the knockout virus exhibited a single-cell infection phenotype. Titration assays confirmed that no infectious budded viruses (BVs) were produced due to the bm40 deletion, while there was no effect on viral DNA replication. Electron microscopy revealed that Bm40 is required for nucleocapsid egress from the nucleus to the cytoplasm, nucleocapsid envelopment to form occlusion-derived viruses (ODVs) and subsequent embedding of ODVs into polyhedra. Confocal microscopy showed that Bm40 was predominantly localized in the nuclei from 48 h post-infection and subsequently condensed on the nuclear membrane and polyhedra at the late phase of infection. Taken together, these results demonstrate that Bm40 plays an essential role in BV production and ODV envelopment in the BmNPV infection cycle.