A Progeny Virus from a Cucumovirus Pseudorecombinant Evolved to Gain the Ability to Accumulate Its RNA-Silencing Suppressor Leading to Systemic Infection in Tobacco
A Progeny Virus from a Cucumovirus Pseudorecombinant Evolved to Gain the Ability to Accumulate Its RNA-Silencing Suppressor Leading to Systemic Infection in Tobacco
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DOI：
10.1094/mpmi-23-3-0332
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发表时间：
2010-03-01
影响因子：
3.5
通讯作者：
Masuta, Chikara
中科院分区：
文献类型：
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作者：
Asaoka, Ryota;Shimura, Hanako;Masuta, Chikara
番茄无精病毒（Tomato aspermy virus，TAV）的两个分离物V-TAV和C-TAV能系统侵染本氏烟草（Nicotiana benthamiana），而只有C-TAV能系统侵染烟草（Nicotiana benthamiana）。烟草这两个株系之间的任何假重组体都不能像C-TAV那样在烟草中有效地系统移动。然而，由V-TAV的RNA 1和3以及C-TAV的RNA 2组成的假重组体（V1 C2 V3）不能全身感染烟草，产生了在V1中具有突变和在C2中具有重组的子代（V1（m）C2（r）V3），使得病毒能够全身移动。为了避免病毒中的进一步突变和重组，我们使用黄瓜花叶病毒RNA 3（Y3）进行后续实验。北方印迹分析表明，编码2b蛋白（2b）的RNA 4A和RNA 5在感染V1（m）C2（r）Y3的烟草中大量积累。在Western印迹中，V1（m）C2（r）Y3实际上比V1 C2 Y3引起更高的2b积累，并且PVX载体的2b过表达使V1 C2 Y3能够在烟草中系统移动，表明2b积累促进病毒系统移动。由于2b的RNA沉默抑制子（RSS）活性被认为参与系统运动，我们比较了两个TAV分离物的2b的RSS活性;在瞬时原生质体测定中，C-TAV 2b在烟草中的活性比V-TAV 2b强。我们的数据还表明，2b和RNA 5通过产生突变/重组病毒和RNA沉默的病毒系统运动，在黄瓜病毒属成员的进化中发挥重要作用。
Two isolates of Tomato aspermy virus (TAV), V-TAV and C-TAV, can systemically infect Nicotiana benthamiana but only C-TAV can move systemically in N. tabacum. Any pseudorecombinants between the two strains could not move systemically in tobacco as efficiently as C-TAV. However, a pseudorecombinant consisting of RNAs 1 and 3 of V-TAV and RNA 2 of C-TAV (V1C2V3), which cannot infect tobacco systemically, generated progeny with a mutation in V1 and a recombination in C2 (V1(m)C2(r)V3), enabling the virus to move systemically. To avoid further mutation and recombination in the virus, we used Cucumber mosaic virus RNA3 (Y3) for subsequent experiments. Northern blot analyses showed that RNA4A, which encodes the 2b protein (2b), and RNA5 abundantly accumulated in V1(m)C2(r)Y3-infected tobacco. V1(m)C2(r)Y3 actually caused higher accumulation of 2b than did V1C2Y3 in Western blots, and overexpression of 2b by the PVX vector enabled V1C2Y3 to move systemically in tobacco, suggesting that 2b accumulation promotes viral systemic movement. Because RNA-silencing suppressor (RSS) activity of 2b was thought to be involved in systemic movement, we compared the RSS activity of 2b for the two TAV isolates; C-TAV 2b had stronger activity than did V-TAV 2b in tobacco in a transient protoplast assay. Our data also demonstrated that 2b and RNA5 play an important role in the evolution of members of genus Cucumovirus by generating mutant/recombinant viruses and viral systemic movement over RNA silencing.