Tomato yellow leaf curl virus (TYLCV) is a DNA virus belonging to the genus Begomovirus. TYLCV replicates using double-stranded DNA intermediates that can become the target of plant transcriptional gene silencing (TGS). Here, we show that the V2 protein of TYLCV can suppress TGS of a transcriptionally silenced green fluorescent protein (GFP) transgene in Nicotiana benthamiana line 16-TGS. Through bisulfite sequencing and chop-PCR, we demonstrated that the TYLCV V2 can reverse GFP transgene silencing by reducing the methylation levels in the 35S promoter sequence. Both AtSN1 and MEA-ISR loci in Arabidopsis thaliana were previously reported to be strongly methylated, and we show that the methylation status of both loci was significantly reduced in TYLCV V2 transgenic Arabidopsis plants. We conclude that TYLCV can efficiently suppress TGS when it infects plants, and its V2 protein is responsible for the TGS suppression activity.Cytosine DNA methylation is a conserved epigenetic silencing mechanism modulating many important biological processes (Bird, 2002;Goll & Bestor, 2005;Zhang et al., 2006) and a defence against biotic stresses such as geminivirus infection (Bisaro, 2006;Raja et al., 2010; Yang et al., 2011). Geminiviruses (family Geminiviridae) are a group of plant-infecting viruses containing circular, singlestranded DNA (ssDNA) genomes packaged into twinned particles (Dry et al., 1993;Fauquet et al., 2008;Zhang et al., 2009;Zhou, 2013). Geminiviruses are transmitted by whiteflies or leafhoppers and can cause significant damage to agronomically important crops (Navot et al., 1991;Hanley-Bowdoin et al., 2004;Rojas et al., 2005). Geminiviruses are classified into four genera (Mastrevirus, Curtovirus, Begomovirus, and Topocuvirus) and replicate in the plant cell nucleus using double-stranded DNA (dsDNA) intermediates as their replication and transcription templates (Pilartz & Jeske, 1992Fauquet et al., 2008).Recent studies have demonstrated that plants employ an RNA-directed methylation strategy to control transcriptional gene silencing (TGS) and use it as a defence mechanism against geminivirus infection (Raja et al., 2008;Rodríguez-Negrete et al., 2009). As a counter defensive measure, geminiviruses express unique proteins to serve as TGS suppressors. For example, the AC2/AL2 proteins encoded by cabbage leaf curl virus and tomato golden mosaic virus, the C2/L2 proteins encoded by beet curly top virus and beet severe curly top virus, and the bC1 protein encoded by tomato yellow leaf curl China betasatellite (TYLCCNB) can decrease DNA methylation and suppress TGS by interfering with the methyl cycle (Buchmann et al., 2009; Yang et al., 2011;Zhang et al., 2011). A recent report indicated that geminivirus replication-associated protein (Rep, also known as C1, AL1 or AC1) could also suppress TGS by reducing the expression of plant DNA methyltransferases (Rodríguez-Negrete et al., 2013).Tomato yellow leaf curl virus (TYLCV) is a member of the genus Begomovirus and contains a single genome component with six open r...

A novel virus, tentatively named Maize Yellow Mosaic Virus (MaYMV), was identified from the field-grown maize plants showing yellow mosaic symptoms on the leaves collected from the Yunnan Province of China by the deep sequencing of small RNAs. The complete 5642 nucleotide (nt)-long genome of the MaYMV shared the highest nucleotide sequence identity (73%) to Maize Yellow Dwarf Virus-RMV. Sequence comparisons and phylogenetic analyses suggested that MaYMV represents a new member of the genus Polerovirus in the family Luteoviridae. Furthermore, the P0 protein encoded by MaYMV was demonstrated to inhibit both local and systemic RNA silencing by co-infiltration assays using transgenic Nicotiana benthamiana line 16c carrying the GFP reporter gene, which further supported the identification of a new polerovirus. The biologically-active cDNA clone of MaYMV was generated by inserting the full-length cDNA of MaYMV into the binary vector pCB301. RT-PCR and Northern blot analyses showed that this clone was systemically infectious upon agro-inoculation into N. benthamiana. Subsequently, 13 different isolates of MaYMV from field-grown maize plants in different geographical locations of Yunnan and Guizhou provinces of China were sequenced. Analyses of their molecular variation indicate that the 3′ half of P3–P5 read-through protein coding region was the most variable, whereas the coat protein- (CP-) and movement protein- (MP-)coding regions were the most conserved.

BackgroundCucumber green mottle mosaic virus (CGMMV), a member of the genus Tobamovirus, can be transmitted by seeds and infects many cucurbit species, causing serious yield losses in cucumber and watermelon plants. In this paper, five serological methods including antigen-coated plate enzyme-linked immunosorbent assay (ACP-ELISA), triple antibody sandwich enzyme-linked immunosorbent assay (TAS-ELISA), Dot-immunobinding assay (DBIA), direct tissue blot immunoassay (DTBIA) and immunocapture reverse transcriptase polymerase chain reaction (IC-RT-PCR) were described for detection and diagnosis of CGMMV.ResultsUsing the purified CGMMV particles as immunogens, six murine monoclonal antibodies (MAbs) were produced. Five serological methods were established using the MAb 4H1 and detection sensitivity was compared using purified preparations and infected-plant tissue extracts. The detection sensitivity of ACP-ELISA was 0.16 ng of purified CGMMV, whereas TAS-ELISA was more sensitive than ACP-ELISA with a minimum detection of 0.04 ng of purified CGMMV. The sensitivities of TAS-ELISA and DBIA were similar for detecting CGMMV in infected-plant tissue extracts, and were four times higher than ACP-ELISA. The IC-RT-PCR was the most sensitive method, which could detect as little as 0.1 pg of purified virus. The detection sensitivity of IC-RT-PCR for CGMMV-infected plant tissues was about 400 times higher than that of TAS-ELISA and DBIA.ConclusionsThe established ACP-ELISA, TAS-ELISA, DBIA and DTBIA are suitable for routine CGMMV detection of large-scale samples in the field survey, while IC-RT-PCR is more sensitive and suitable for acquiring information about the viral genome.