Tomato Chlorotic Mottle Virus Is a Target of RNA Silencing but the Presence of Specific Short Interfering RNAs Does Not Guarantee Resistance in Transgenic Plants

Abstract

Tomato chlorotic mottle virus (ToCMoV) is a begomovirus found widespread in tomato fields in Brazil.ToCMoV isolate BA-Se1 (ToCMoV-[BA-Se1]) was shown to trigger the plant RNA silencing surveillance in different host plants and, coinciding with a decrease in viral DNA levels, small interfering RNAs (siRNAs) specific to ToCMoV-[BA-Se1] accumulated in infected plants. Although not homogeneously distributed, the siRNA population in both infected Nicotiana benthamiana and tomato plants represented the entire DNA-A and DNA-B genomes. We determined that in N. benthamiana, the primary targets corresponded to the 5 end of AC1 and the embedded AC4, the intergenic region and 5 end of AV1 and overlapping central part of AC5. Subsequently, transgenic N. benthamiana plants were generated that were preprogrammed to express double-stranded RNA corresponding to this most targeted portion of the virus genome by using an intron-hairpin construct. These plants were shown to indeed produce ToCMoVspecific siRNAs. When challenge inoculated, most transgenic lines showed significant delays in symptom development, and two lines had immune plants. Interestingly, the levels of transgene-produced siRNAs were similar in resistant and susceptible siblings of the same line. This indicates that, in contrast to RNA viruses, the mere presence of transgene siRNAs corresponding to DNA virus sequences does not guarantee virus resistance and that other factors may play a role in determining RNA-mediated resistance to DNA viruses.

RNA Silencing against Geminivirus: Complementary Action of Posttranscriptional Gene Silencing and Transcriptional Gene Silencing in Host Recovery

“…We have also corroborated that the virus-associated siRNA populations are related to the entire genome although their concentrations are not homogeneous. Again, it seems that each virus or virus-host combination might present a particular pattern (14,36,45). For PepGMV, zones corresponding to transcripts of the Rep, TrAP, REn, and MP genes and both IRs seem to be preferential targets (hot spots) for a silencing mechanism in highly symptomatic tissue (Fig.…”

“…Recent reports have shown that geminivirus-infected plants can recover or show reduction of symptoms (1,12,45). Such processes have been correlated to RNA silencing mechanisms.…”

Tomato yellow leaf curl virus resistance by Ty-1 involves increased cytosine methylation of viral genomes and is compromised by cucumber mosaic virus infection

“…Although geminiviruses harbor a single-stranded DNA genome, they also are subject to transcriptional and posttranscriptional RNA silencing, as observed by the synthesis of geminivirus-specific siRNAs, (sRNA-directed) viral DNA methylation, and posttranscriptional gene silencing of protein-coding genes (14)(15)(16)(17). However, antiviral RNAi defense against geminiviruses is suggested to mostly rely on a methylation-based defense, a process that involves the action of siRNA-directed methylation pathway component Ago4 (18,19).…”

Application of Virus-Derived Small Interfering RNAs (vsiRNAs) in Rice Viruses with Insect Vectors, Especially <em>Rice Grassy Stunt Virus</em>

“…Genetic Engineering is most important technology for determining the resistance to field crops, whether it is found or not used by virus-derived genes that lead to the pathogen-derived resistance [79]. RNA Silencing is one of the most widely used methods to check the resistance in plants against the RNA viruses which is approximately used for more than 15 years [14, 64,74].…”