National Institute for Biotechnology and Genetic Engineering, Faisalabad, Pakistan.
Mol Plant Microbe Interact. 2011 Aug;24(8):973-83. doi: 10.1094/MPMI-01-11-0001.
Begomoviruses (family Geminiviridae) are single-stranded DNA viruses transmitted by the whitefly Bemisia tabaci. Many economically important diseases in crops are caused by begomoviruses, particularly in tropical and subtropical environments. These include the betasatellite-associated begomoviruses causing cotton leaf curl disease (CLCuD) that causes significant losses to a mainstay of the economy of Pakistan, cotton. RNA interference (RNAi) or gene silencing is a natural defense response of plants against invading viruses. In counter-defense, viruses encode suppressors of gene silencing that allow them to effectively invade plants. Here, we have analyzed the ability of the begomovirus Cotton leaf curl Multan virus (CLCuMV) and its associated betasatellite, Cotton leaf curl Multan β-satellite (CLCuMB) which, together, cause CLCuD, and the nonessential alphasatellite (Cotton leaf curl Multan alphasatellite [CLCuMA]) for their ability to suppress gene silencing in Nicotiana benthamiana. The results showed that CLCuMV by itself was unable to efficiently block silencing. However, in the presence of the betasatellite, gene silencing was entirely suppressed. Silencing was not affected in any way when infections included CLCuMA, although the alphasatellite was, for the first time, shown to be a target of RNA silencing, inducing the production in planta of specific small interfering RNAs, the effectors of silencing. Subsequently, using a quantitative real-time polymerase chain reaction assay and Northern blot analysis, the ability of all proteins encoded by CLCuMV and CLCuMB were assessed for their ability to suppress RNAi and the relative strengths of their suppression activity were compared. The analysis showed that the V2, C2, C4, and βC1 proteins exhibited suppressor activity, with the V2 showing the strongest activity. In addition, V2, C4, and βC1 were examined for their ability to bind RNA and shown to have distinct specificities. Although each of these proteins has, for other begomoviruses or betasatellites, been previously shown to have suppressor activity, this is the first time all proteins encoded by a geminiviruses (or begomovirus-betasatellite complex) have been examined and also the first for which four separate suppressors have been identified.
双生病毒（家族 Geminiviridae）是通过烟粉虱 Bemisia tabaci 传播的单链 DNA 病毒。许多在农作物中造成经济损失的重要疾病都是由双生病毒引起的，特别是在热带和亚热带环境中。其中包括与卫星 β 相伴的贝塔卫星引起的棉花卷曲叶病（CLCuD），这对巴基斯坦经济支柱棉花造成了重大损失。RNA 干扰（RNAi）或基因沉默是植物抵御入侵病毒的一种自然防御反应。在反防御中，病毒编码基因沉默抑制剂，使它们能够有效地入侵植物。在这里，我们分析了引起 CLCuD 的双生病毒棉花卷曲 Multan 病毒（CLCuMV）及其相关的卫星β-Cotton leaf curl Multan β-卫星（CLCuMB）以及非必需的α卫星（棉花卷曲 Multan 阿尔法卫星[CLCuMA]）抑制 Nicotiana benthamiana 中基因沉默的能力。结果表明，CLCuMV 本身不能有效地阻断沉默。然而，在存在卫星β的情况下，基因沉默完全被抑制。当感染包括 CLCuMA 时，沉默没有受到任何影响，尽管首次发现α卫星是 RNA 沉默的靶标，在植物体内诱导产生特定的小干扰 RNA，即沉默的效应物。随后，使用定量实时聚合酶链反应测定和 Northern blot 分析，评估了 CLCuMV 和 CLCuMB 编码的所有蛋白质抑制 RNAi 的能力，并比较了它们抑制活性的相对强度。分析表明，V2、C2、C4 和βC1 蛋白表现出抑制活性，其中 V2 表现出最强的活性。此外，还检测了 V2、C4 和βC1 结合 RNA 的能力，并显示出它们具有不同的特异性。尽管以前已经证明其他双生病毒或卫星β中的这些蛋白都具有抑制活性，但这是首次检查由双生病毒（或双生病毒-卫星β复合物）编码的所有蛋白，也是首次鉴定出四个独立的抑制蛋白。
