Identification of the virulence factors and suppressors of posttranscriptional gene silencing encoded by Ageratum yellow vein virus, a monopartite begomovirus

Ageratum yellow vein disease (AYVD) is caused by the association of a Tomato leaf curl Java betasatellite [Indonesia:Indonesia 1:2003] (ToLCJB-[ID:ID1:03]) with a begomovirus component. Our previous results demonstrated that ToLCJB-[ID:ID:03] is essential for induction of leaf curl symptoms in plants and transgene expression of its βC1 gene in Nicotiana benthamiana plants induces virus-like symptoms. Here we show that Ageratum yellow vein virus-Indonesia [Indonesia: Tomato] (AYVV-ID[ID:Tom]) alone could systemically infect the plants and induced upward leaf curl symptoms. ToLCJB-[ID:ID1:03] was required, in addition to AYVV-ID[ID:Tom], for induction of severe downward leaf curl disease in N. benthamiana plants. However, DNAβ01fsβC1, which encompasses a frameshift mutation, did not induce severe symptoms in N. benthamiana when co-inoculated with AYVV-ID[ID:Tom]. The infectivity analysis of AYVV-ID[ID:Tom] and its associated betasatellite encoded genes using Potato virus X (PVX) vector were carried out in N. benthamiana, indicate that the V2 and βC1 genes are symptom determinants. We have identified the DNA encoded V2 and its betasatellite, ToLCJB-[ID:ID1:03], encoded βC1 proteins as efficient silencing suppressors of posttranscriptional gene silencing (PTGS) by using an Agrobacterium co-infiltration or heterologous PVX vector assays. However, the results also showed weak suppression of gene silencing activities for C2 and C4 induced by GFP and mRNA associated with GFP was detected. Furthermore, confocal imaging analysis of ToLCJB-[ID:ID1:03] βC1 in the epidermal cells of N. benthamiana shows that this protein is accumulated towards the periphery of the cell and around the nucleus, however, V2 accumulated in the cell cytoplasm, C4 associated with plasma membrane and C2 exclusively targeted into nucleus. In this study, we identified as many as four distinct suppressors of RNA silencing encoded by AYVV-ID[ID:Tom] and its cognate betasatellite in the family Geminiviridae, counteracting innate antiviral response.