Suppressors of RNA Silencing Encoded by the Components of the Cotton Leaf Curl Begomovirus-BetaSatellite Complex

Abstract

Begomoviruses (family Geminiviridae) are single-stranded DNA viruses transmitted by the whitefly Bemisia tabaci. Many economically important diseases in crops are caused by begomoviruses, particularly in tropical and subtropical environments. These include the betasatellite-associated begomoviruses causing cotton leaf curl disease (CLCuD) that causes significant losses to a mainstay of the economy of Pakistan, cotton. RNA interference (RNAi) or gene silencing is a natural defense response of plants against invading viruses. In counter-defense, viruses encode suppressors of gene silencing that allow them to effectively invade plants. Here, we have analyzed the ability of the begomovirus Cotton leaf curl Multan virus (CLCuMV) and its associated betasatellite, Cotton leaf curl Multan β-satellite (CLCuMB) which, together, cause CLCuD, and the nonessential alphasatellite (Cotton leaf curl Multan alphasatellite [CLCuMA]) for their ability to suppress gene silencing in Nicotiana benthamiana. The results showed that CLCuMV by itself was unable to efficiently block silencing. However, in the presence of the betasatellite, gene silencing was entirely suppressed. Silencing was not affected in any way when infections included CLCuMA, although the alphasatellite was, for the first time, shown to be a target of RNA silencing, inducing the production in planta of specific small interfering RNAs, the effectors of silencing. Subsequently, using a quantitative real-time polymerase chain reaction assay and Northern blot analysis, the ability of all proteins encoded by CLCuMV and CLCuMB were assessed for their ability to suppress RNAi and the relative strengths of their suppression activity were compared. The analysis showed that the V2, C2, C4, and βC1 proteins exhibited suppressor activity, with the V2 showing the strongest activity. In addition, V2, C4, and βC1 were examined for their ability to bind RNA and shown to have distinct specificities. Although each of these proteins has, for other begomoviruses or betasatellites, been previously shown to have suppressor activity, this is the first time all proteins encoded by a geminiviruses (or begomovirus-betasatellite complex) have been examined and also the first for which four separate suppressors have been identified.

Dominance of resistance-breaking cotton leaf curl Burewala virus (CLCuBuV) in northwestern India

“…The different functions attributed to the C2 protein, such as transcription activation [22,48,55,57], suppression of post-transcriptional gene silencing (PTGS) [3,62], suppression of cell death [3,7] and interactions with a variety of host factors have been reviewed by Amin et al [3]. Our preliminary studies show that CLCuBuV-C2-2S cannot transactivate the V1 gene (Meera et al, unpublished data), and the same was observed by Amin et al [3] for CLCuMuV.…”

“…Our preliminary studies show that CLCuBuV-C2-2S cannot transactivate the V1 gene (Meera et al, unpublished data), and the same was observed by Amin et al [3] for CLCuMuV. Among the different suppressors, V2, C2, C4 and bC1, encoded by CLCuMuV, C2 has weak activity according to Amin et al [3].…”

Association of Tomato Yellow Leaf Curl Virus and Chili Leaf Curl Virus with Leaf Curl Disease of Radish and the Synergistic Interaction on Nicotiana benthamiana

“…These proteins are referred to as suppressors of gene silencing. Usually viruses encode multiple suppressors that can silence the RNAi pathway at multiple places (Vanitharani et al, 2004;Amin et al, 2011). In synergistic interactions, these suppressors of gene silencing play an important role.…”

“…A bipartite genome is the genome that consists of both components together, referred to as DNA A and DNA B. Begomoviruses belonging to the Old World (OW), in contrast, have one or two genomic components, monopartite or bipartite genomes, respectively. Betasatellites are also known to encode silencing suppressors (Amin et al, 2011). Betasatellites are required for the infectivity of some viruses (Briddon and Stanley, 2006).…”

“…However, it has been shown that the suppressor activity of the geminiviral protein may play an important role in this. Different geminiviral proteins from different viruses may act as suppressors of gene silencing; their strength and mode of action may also differ like TrAP, C4, and V2 (Vanitharani et al, 2004;Amin et al, 2011). Synergism may lead to altered movement of the virus (Alves-Júnior et al, 2009).…”