title: Field Trial and Molecular Characterization of RNAi-Transgenic Tomato Plants That Exhibit Resistance to Tomato Yellow Leaf Curl Geminivirus.
abstract: RNA interference (RNAi) is a widely used approach to generate virus-resistant transgenic crops. However, issues of agricultural importance like the long-term durability of RNAi-mediated resistance under field conditions and the potential side effects provoked in the plant by the stable RNAi expression remain poorly investigated. Here, we performed field trials and molecular characterization studies of two homozygous transgenic tomato lines, with different selection markers, expressing an intron-hairpin RNA cognate to the Tomato yellow leaf curl virus (TYLCV) C1 gene. The tested F6 and F4 progenies of the respective kanamycin- and basta-resistant plants exhibited unchanged field resistance to TYLCV and stably expressed the transgene-derived short interfering RNA (siRNAs) to represent 6 to 8% of the total plant small RNAs. This value outnumbered the average percentage of viral siRNAs in the nontransformed plants exposed to TYLCV-infested whiteflies. As a result of the RNAi transgene expression, a common set of up- and downregulated genes was revealed in the transcriptome profile of the plants selected from either of the two transgenic events. A previously unidentified geminivirus causing no symptoms of viral disease was detected in some of the transgenic plants. The novel virus acquired V1 and V2 genes from TYLCV and C1, C2, C3, and C4 genes from a distantly related geminivirus and, thereby, it could evade the repressive sequence-specific action of transgene-derived siRNAs. Our findings shed light on the mechanisms of siRNA-directed antiviral silencing in transgenic plants and highlight the applicability limitations of this technology as it may alter the transcriptional pattern of nontarget genes.
authors.affiliation: 1 Center for Genetic Engineering and Biotechnology, calle 31 entre 158 y 190, Cubanacan Playa, Apdo 6162, Habana 10600, Cuba;
authors.forename: Alejandro
authors.initials: A
authors.lastname: Fuentes
authors.affiliation: 1 Center for Genetic Engineering and Biotechnology, calle 31 entre 158 y 190, Cubanacan Playa, Apdo 6162, Habana 10600, Cuba;
authors.forename: Natacha
authors.initials: N
authors.lastname: Carlos
authors.affiliation: 1 Center for Genetic Engineering and Biotechnology, calle 31 entre 158 y 190, Cubanacan Playa, Apdo 6162, Habana 10600, Cuba;
authors.forename: Yoslaine
authors.initials: Y
authors.lastname: Ruiz
authors.affiliation: 1 Center for Genetic Engineering and Biotechnology, calle 31 entre 158 y 190, Cubanacan Playa, Apdo 6162, Habana 10600, Cuba;
authors.forename: Danay
authors.initials: D
authors.lastname: Callard
authors.affiliation: 1 Center for Genetic Engineering and Biotechnology, calle 31 entre 158 y 190, Cubanacan Playa, Apdo 6162, Habana 10600, Cuba;
authors.forename: Yadira
authors.initials: Y
authors.lastname: Sánchez
authors.affiliation: 1 Center for Genetic Engineering and Biotechnology, calle 31 entre 158 y 190, Cubanacan Playa, Apdo 6162, Habana 10600, Cuba;
authors.forename: María Elena
authors.initials: ME
authors.lastname: Ochagavía
authors.affiliation: 2 University of Basel, Department of Environmental Sciences, Botany, Hebelstrasse 1, 4056 Basel, Switzerland;|3 FASTERIS SA, Ch. Du Pont-du-Centenaire 109, 1228 Plan-les-Ouates, Switzerland; and.
authors.forename: Jonathan
authors.initials: J
authors.lastname: Seguin
authors.affiliation: 2 University of Basel, Department of Environmental Sciences, Botany, Hebelstrasse 1, 4056 Basel, Switzerland;
authors.forename: Nachelli
authors.initials: N
authors.lastname: Malpica-López
authors.affiliation: 2 University of Basel, Department of Environmental Sciences, Botany, Hebelstrasse 1, 4056 Basel, Switzerland;
authors.forename: Thomas
authors.initials: T
authors.lastname: Hohn
authors.affiliation: 4 Functional Genomics Center ETH Zurich, University of Zurich, Winterthurerstrasse 190/Y32 H80, 8057 Zurich, Switzerland.
authors.forename: Maria Rita
authors.initials: MR
authors.lastname: Lecca
authors.affiliation: 1 Center for Genetic Engineering and Biotechnology, calle 31 entre 158 y 190, Cubanacan Playa, Apdo 6162, Habana 10600, Cuba;
authors.forename: Rosabel
authors.initials: R
authors.lastname: Pérez
authors.affiliation: 1 Center for Genetic Engineering and Biotechnology, calle 31 entre 158 y 190, Cubanacan Playa, Apdo 6162, Habana 10600, Cuba;
authors.forename: Vivian
authors.initials: V
authors.lastname: Doreste
authors.affiliation: 4 Functional Genomics Center ETH Zurich, University of Zurich, Winterthurerstrasse 190/Y32 H80, 8057 Zurich, Switzerland.
authors.forename: Hubert
authors.initials: H
authors.lastname: Rehrauer
authors.affiliation: 3 FASTERIS SA, Ch. Du Pont-du-Centenaire 109, 1228 Plan-les-Ouates, Switzerland; and.
authors.forename: Laurent
authors.initials: L
authors.lastname: Farinelli
authors.affiliation: 1 Center for Genetic Engineering and Biotechnology, calle 31 entre 158 y 190, Cubanacan Playa, Apdo 6162, Habana 10600, Cuba;
authors.forename: Merardo
authors.initials: M
authors.lastname: Pujol
authors.affiliation: 2 University of Basel, Department of Environmental Sciences, Botany, Hebelstrasse 1, 4056 Basel, Switzerland;
authors.forename: Mikhail M
authors.initials: MM
authors.lastname: Pooggin
mesh_terms: D017791:Geminiviridae / Q000502:physiology*; D018506:Gene Expression Regulation, Plant; D020022:Genetic Predisposition to Disease; D018551:Solanum lycopersicum / Q000235:genetics* / Q000502:physiology*; D008969:Molecular Sequence Data; D010935:Plant Diseases / Q000235:genetics / Q000821:virology*; D030821:Plants, Genetically Modified; D034622:RNA Interference; D034741:RNA, Small Interfering; D059467:Transcriptome
chemical_list: D034741:RNA, Small Interfering