Abstract
RNA silencing is an important defense mechanism in plants, yet several plant viruses encode proteins that suppress this mechanism. In this study, the genome of the Olive mild mosaic virus (OMMV) was screened for silencing suppressors. The full OMMV cDNA and 5 OMMV open reading frames (ORFs) were cloned into the Gateway binary vector pK7WG2, transformed into Agrobacterium tumefaciens, and agroinfiltrated into N. benthamiana 16C plants. CP and p6 showed suppressor activity, with CP showing significantly higher activity than p6, yet activity that was lower than the full OMMV, suggesting a complementary action of CP and p6. These viral suppressors were then used to induce OMMV resistance in plants based on RNA silencing. Two hairpin constructs targeting each suppressor were agroinfiltrated in N. benthamiana plants, which were then inoculated with OMMV RNA. When silencing of both suppressors was achieved, a significant reduction in viral accumulation and symptom attenuation was observed as compared to those of the controls, as well as to when each construct was used alone, proving them to be effective against OMMV infection. This is the first time that a silencing suppressor was found in a necrovirus, and that two independent proteins act as silencing suppressors in a virus member of the Tombusviridae family.
Highlights
RNA silencing is a gene inactivation mechanism identified in most eukaryotes that is involved in several biological processes such as regulating endogenous gene expression, the maintenance of genome stability, and defense against viruses [1,2].Amongst the several strategies plants have developed to counter virus infections, RNA silencing is one of the most important [1,3]
10, x potential Olive mild mosaic virus (OMMV) RNA silencing suppressors, all of the proteins encoded by6 the of 14 genome were tested for their ability to suppress silencing by using a green fluorescent protein (GFP) reporter gene in plant tissues
Members of the Tombusviridae family present a high diversity of viral suppressors
Summary
Amongst the several strategies plants have developed to counter virus infections, RNA silencing is one of the most important [1,3]. Plant virus infections are associated with the accumulation of these virus-specific siRNAs. The cleavage is accomplished by members of the Argonaute protein family (AGOs) [4] that recruit siRNA and associated proteins to form the RNA-induced silencing complex (RISC). The cleavage is accomplished by members of the Argonaute protein family (AGOs) [4] that recruit siRNA and associated proteins to form the RNA-induced silencing complex (RISC) This complex possesses ribonuclease activity and is guided by the single-stranded siRNAs to Viruses 2018, 10, 416; doi:10.3390/v10080416 www.mdpi.com/journal/viruses
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