We investigated the genetic pathway in Arabidopsis thaliana targeted during infection by cucumber mosaic virus (CMV) 2b protein, known to suppress non-cell-autonomous transgene silencing and salicylic acid (SA)-mediated virus resistance. We show that 2b expressed from the CMV genome drastically reduced the accumulation of 21-, 22-, and 24-nucleotide classes of viral small interfering RNAs (siRNAs) produced by Dicer-like4 (DCL4), DCL2, and DCL3, respectively. The defect of a CMV 2b-deletion mutant (CMV-Delta2b) in plant infection was efficiently rescued in Arabidopsis mutants producing neither 21- nor 22-nucleotide viral siRNAs. Since genetic analysis further identifies a unique antiviral role for DCL3 upstream of DCL4, our data indicate that inhibition of the accumulation of distinct viral siRNAs plays a key role in 2b suppression of antiviral silencing. Strikingly, disease symptoms caused by CMV-Delta2b in Arabidopsis mutants defective in antiviral silencing were as severe as those caused by CMV, demonstrating an indirect role for the silencing suppressor activity in virus virulence. We found that production of CMV siRNAs without 2b interference depended largely on RNA-dependent RNA polymerase 1 (RDR1) inducible by SA. Given the known role of RDR6-dependent transgene siRNAs in non-cell-autonomous silencing, our results suggest a model in which 2b inhibits the production of RDR1-dependent viral siRNAs that confer SA-dependent virus resistance by directing non-cell-autonomous antiviral silencing.

译文

我们研究了在黄瓜花叶病毒 (CMV) 2b蛋白感染期间针对拟南芥的遗传途径,该蛋白已知可抑制非细胞自主转基因沉默和水杨酸 (SA) 介导的病毒抗性。我们显示,从CMV基因组表达的2b分别大大减少了由Dicer-like4 (DCL4),DCL2和DCL3产生的21、22和24核苷酸类病毒小干扰rna (sirna) 的积累。在既不产生21个或22个核苷酸的病毒sirna的拟南芥突变体中,有效地挽救了植物感染中CMV 2b缺失突变体 (CMV-Delta2b) 的缺陷。由于遗传分析进一步确定了DCL4上游DCL3的独特抗病毒作用,因此我们的数据表明,抑制不同病毒sirna的积累在2b抑制抗病毒沉默中起着关键作用。令人惊讶的是,在抗病毒沉默中缺陷的拟南芥突变体中由CMV-Delta2b引起的疾病症状与由CMV引起的症状一样严重,表明沉默抑制活性在病毒毒力中的间接作用。我们发现,没有2b干扰的CMV sirna的产生在很大程度上取决于SA诱导的RNA依赖性RNA聚合酶1 (RDR1)。鉴于RDR6-dependent转基因sirna在非细胞自主沉默中的已知作用,我们的结果提出了一种模型,其中2b通过指导非细胞自主抗病毒沉默抑制赋予SA依赖性病毒抗性的RDR1-dependent病毒sirna的产生。

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