College of Plant Protection and Shaanxi Key Laboratory of Molecular Biology for Agriculture, Northwest A & F University, Yangling, Shaanxi 712100, PR China.
Virus Res. 2011 Feb;155(2):423-32. doi: 10.1016/j.virusres.2010.11.011. Epub 2010 Dec 7.
The p20 protein encoded by the tombusvirus, Cucumber necrosis virus has previously been shown to be involved in host pathogenicity and shares sequence similarity with the Tomato bushy stunt virus p19 suppressor of silencing. Using a virus-induced gene silencing (VIGS) assay, we show that p20 is a viral suppressor of RNA silencing (VSR) in infected plants. In addition, a CNV p20-knockout mutant showed a decline in viral RNA accumulation in infected plants, consistent with the role of p20 in suppression of RNA silencing. However, unexpectedly, all GFP transgenic plants co-infiltrated with p20 and GFP displayed RNA silencing using an Agrobacterium-mediated silencing assay. Detailed RNA analysis of GFP mRNA levels in p20 agro-infiltrated plants revealed that p20 did initially display suppressor activity but this was rapidly overcome by RNA silencing. p20 expression levels in agro-infiltrated plants were shown to be approximately 50-fold lower than that of the TBSV p19 silencing suppressor, consistent with the notion that p20 dosage levels are not sufficient to suppress RNA silencing in the Agrobacterium-mediated system. Our results suggest that a viral-based VIGS assay may be required for identifying VSRs encoded by some plant viruses. Based on bioinformatics studies the mechanism of suppression of silencing by p20 is predicted to be similar to that of the TBSV p19 suppressor.
由 Tospovirus 编码的 p20 蛋白先前被证明参与宿主致病性，并与番茄丛矮病毒的 p19 沉默抑制子具有序列相似性。使用病毒诱导的基因沉默（VIGS）测定，我们表明 p20 是感染植物中 RNA 沉默（RS）的病毒抑制剂。此外，CNV p20 敲除突变体显示感染植物中病毒 RNA 积累下降，与 p20 在抑制 RNA 沉默中的作用一致。然而，出乎意料的是，所有共浸润 p20 和 GFP 的 GFP 转基因植物均显示出通过农杆菌介导的沉默测定的 RNA 沉默。对 p20 agro 浸润植物中 GFP mRNA 水平的详细 RNA 分析表明，p20 最初确实显示出抑制活性，但很快被 RNA 沉默所克服。在 agro 浸润植物中 p20 的表达水平比 TBSV p19 沉默抑制剂低约 50 倍，这与 p20 剂量水平不足以抑制农杆菌介导系统中 RNA 沉默的观点一致。我们的结果表明，可能需要基于病毒的 VIGS 测定来鉴定某些植物病毒编码的 VSR。基于生物信息学研究，p20 沉默抑制的机制预测与 TBSV p19 抑制剂相似。
