Storms M M, Kormelink R, Peters D, Van Lent J W, Goldbach R W
Department of Virology, Wageningen Agricultural University, Wageningen, The Netherlands.
Virology. 1995 Dec 20;214(2):485-93. doi: 10.1006/viro.1995.0059.
The expression and subcellular location of the 33.6-kDa nonstructural protein NSm of tomato spotted wilt virus (TSWV) was analyzed in Nicotiana rustica plants and protoplasts as a function of time. Immunofluorescent studies in protoplasts isolated from TSWV-infected N. rustica leaves showed that this protein could first be detected close to the periphery of the cell, near the plasmamembrane, and later in tubular structures emerging from the cell surface. In situ, these tubules appeared specifically in the plasmodesmata, suggesting their involvement in cell-to-cell movement of the virus during systemic infection. In protoplasts transfected with an expression vector containing the NSm gene, similar tubules were formed, indicating that NSm has the ability to form these structures in the absence of other virus-specific components. To test whether plant-specific components were involved in tubule formation, the NSm gene was also expressed in a heterologous expression system, i.e., insect cells. Spodoptera frugiperda and Trichoplusia ni cells were infected with a recombinant baculovirus expressing the NSm-gene (AcNPV/NSm). The efficient formation of NSm-containing tubules emerging from the surface of both cell types indicate that no plant-specific cell structures or proteins are involved in their development.
以时间为函数，分析了番茄斑萎病毒（TSWV）33.6 kDa非结构蛋白NSm在烟草植株和原生质体中的表达及亚细胞定位。对从感染TSWV的烟草叶片中分离出的原生质体进行免疫荧光研究表明，该蛋白最初可在靠近细胞周边、质膜附近检测到，随后出现在从细胞表面伸出的管状结构中。在原位，这些小管特别出现在胞间连丝中，表明它们在系统感染期间参与病毒的细胞间移动。在用含有NSm基因的表达载体转染的原生质体中，形成了类似的小管，这表明NSm在没有其他病毒特异性成分的情况下有能力形成这些结构。为了测试植物特异性成分是否参与小管形成，NSm基因也在异源表达系统即昆虫细胞中表达。用表达NSm基因的重组杆状病毒（AcNPV/NSm）感染草地贪夜蛾和粉纹夜蛾细胞。从两种细胞类型表面出现的含NSm小管的有效形成表明，其形成过程不涉及植物特异性细胞结构或蛋白质。
