Tomato fruit as a model for tissue-specific gene silencing in crop plants.
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ABSTRACT:
SUBMITTER: Feder A
PROVIDER: S-EPMC7459100 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:During the growing season of 2021-2022, a total of 145 symptomatic tomato leaf and fruit samples were collected from different locations in Riyadh Region, Saudi Arabia, showing a moderate-to-severe mosaic with dark green wrinkling, blistering, narrowing, and deformation with necrosis spot on tomato leaves, while irregular brown necrotic lesions, deformation, and yellowing spots rendering the fruits non-marketable were observed on tomato fruits. These samples were tested serologically against important tomato viruses using enzyme-linked immunosorbent assay (ELISA), and the obtained results showed that 52.4% of symptomatic tomato samples were found positive for Tomato brown rugose fruit virus (ToBRFV), wherein 12 out of 76 samples were singly infected; however, 64 out of 145 had mixed infection. A sample with a single infection of ToBRFV was used for mechanical inoculation into a range of different host plants; symptoms were observed weekly, and the presence of the ToBRFV was confirmed by ELISA and reverse transcription-polymerase chain reaction (RT-PCR). A total RNA was extracted from selected ELISA-positive samples, and RT-PCR was carried out using specific primers F-3666 and R-4718, which amplified a fragment of 1052 bp. RT-PCR products were sequenced in both directions, and partial genome nucleotide sequences were submitted to GenBank under the following accession numbers: MZ130501, MZ130502, and MZ130503. BLAST analysis of Saudi isolates of ToBRFV showed that the sequence shared nucleotide identities (99-99.5%) among them and 99-100% identity with ToBRFV isolates in different countries. A ToBRFV isolate (MZ130503) was selected for mechanical inoculation and to evaluate symptom severity responses of 13 commonly grown tomato cultivars in Saudi Arabia. All of the tomato cultivars showed a wide range of symptoms. The disease severity index of the tested cultivars ranged between 52% and 96%. The importance ToBRFV disease severity and its expanding host range due to its resistance breaking ability was discussed.
| S-EPMC9692878 | biostudies-literature
Project description:The capacity to assimilate carbon and nitrogen, to transport the resultant sugars and amino acids to sink tissues, and to convert the incoming sugars and amino acids into storage compounds in the sink tissues, are key determinants of crop yield. Given that all of these processes have the potential to co-limit growth, multiple genetic interventions in source and sink tissues, plus transport processes may be necessary to reach the full yield potential of a crop. We used biolistic combinatorial co-transformation (up to 20 transgenes) for increasing C and N flows with the purpose of increasing tomato fruit yield. We observed an increased fruit yield of up to 23%. To better explore the reconfiguration of metabolic networks in these transformants, we generated a dataset encompassing physiological parameters, gene expression and metabolite profiling on plants grown under glasshouse or polytunnel conditions. A Sparse Partial Least Squares regression model was able to explain the combination of genes that contributed to increased fruit yield. This combinatorial study of multiple transgenes targeting primary metabolism thus offers opportunities to probe the genetic basis of metabolic and phenotypic variation, providing insight into the difficulties in choosing the correct combination of targets for engineering increased fruit yield.
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Project description:Key messageSix novel fruit weight QTLs were identified in tomato using multiple bi-parental populations developed from ancestral accessions. Beneficial alleles at these loci arose in semi-domesticated subpopulations and were likely left behind. This study paves the way to introgress these alleles into breeding programs. The size and weight of edible organs have been strongly selected during crop domestication. Concurrently, human have also focused on nutritional and cultural characteristics of fruits and vegetables, at times countering selective pressures on beneficial size and weight alleles. Therefore, it is likely that novel improvement alleles for organ weight still segregate in ancestral germplasm. To date, five domestication and diversification genes affecting tomato fruit weight have been identified, yet the genetic basis for increases in weight has not been fully accounted for. We found that fruit weight increased gradually during domestication and diversification, and semi-domesticated subpopulations featured high phenotypic and nucleotide diversity. Columella and septum fruit tissues were proportionally increased, suggesting targeted selection. We developed twenty-one F2 populations with parents fixed for the known fruit weight genes, corresponding to putative key transitions from wild to fully domesticated tomatoes. These parents also showed differences in fruit weight attributes as well as the developmental timing of size increase. A subset of populations was targeted for QTL-seq, leading to the identification of six uncloned fruit weight QTLs. Three QTLs, located on chromosomes 1, 2 and 3, were subsequently validated by progeny testing. By exploring the segregation of the known fruit weight genes and the identified QTLs, we estimated that most beneficial alleles in the newly identified loci arose in semi-domesticated subpopulations from South America and were not likely transmitted to fully domesticated landraces. Therefore, these alleles could be incorporated into breeding programs using the germplasm and genetic resources identified in this study.
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Project description:Parthenocarpy is potentially a desirable trait for many commercially grown fruits if undesirable changes to structure, flavour, or nutrition can be avoided. Parthenocarpic transgenic tomato plants (cv MicroTom) were obtained by the regulation of genes for auxin synthesis (iaaM) or responsiveness (rolB) driven by DefH9 or the INNER NO OUTER (INO) promoter from Arabidopsis thaliana. Fruits at a breaker stage were analysed at a transcriptomic and metabolomic level using microarrays, real-time reverse transcription-polymerase chain reaction (RT-PCR) and a Pegasus III TOF (time of flight) mass spectrometer. Although differences were observed in the shape of fully ripe fruits, no clear correlation could be made between the number of seeds, transgene, and fruit size. Expression of auxin synthesis or responsiveness genes by both of these promoters produced seedless parthenocarpic fruits. Eighty-three percent of the genes measured showed no significant differences in expression due to parthenocarpy. The remaining 17% with significant variation (P <0.05) (1748 genes) were studied by assigning a predicted function (when known) based on BLAST to the TAIR database. Among them several genes belong to cell wall, hormone metabolism and response (auxin in particular), and metabolism of sugars and lipids. Up-regulation of lipid transfer proteins and differential expression of several indole-3-acetic acid (IAA)- and ethylene-associated genes were observed in transgenic parthenocarpic fruits. Despite differences in several fatty acids, amino acids, and other metabolites, the fundamental metabolic profile remains unchanged. This work showed that parthenocarpy with ovule-specific alteration of auxin synthesis or response driven by the INO promoter could be effectively applied where such changes are commercially desirable.
| S-EPMC2736898 | biostudies-literature
Project description:The tobamovirus tomato brown rugose fruit virus (ToBRFV), a major threat to tomato production worldwide, has recently been documented in mixed infections with the potexvirus pepino mosaic virus (PepMV) CH2 strain in traded tomatoes in Israel. A study of greenhouse tomato plants in Israel revealed severe new viral disease symptoms including open unripe fruits and yellow patched leaves. PepMV was only detected in mixed infections with ToBRFV in all 104 tested sites, using serological and molecular analyses. Six PepMV isolates were identified, all had predicted amino acids characteristic of CH2 mild strains excluding an isoleucine at amino acid position 995 of the replicase. High-throughput sequencing of viral RNA extracted from four selected symptomatic plants showed solely the ToBRFV and PepMV, with total aligned read ratios of 40.61% and 11.73%, respectively, indicating prevalence of the viruses. Analyses of interactions between the co-infecting viruses by sequential and mixed viral inoculations of tomato plants, at various temperatures, showed a prominent increase in PepMV titers in ToBRFV pre-inoculated plants and in mixed-infected plants at 18-25 °C, compared to PepMV-single inoculations, as analyzed by Western blot and quantitative RT-PCR tests. These results suggest that Israeli mild PepMV isolate infections, preceded by ToBRFV, could induce symptoms characteristic of PepMV aggressive strains.
| S-EPMC7472245 | biostudies-literature
Project description:Parthenocarpy is the development of an ovary into a seedless fruit without pollination. The ubiquitous downregulation of SlIAA9 induces not only parthenocarpic fruit formation but also an abnormal vegetative phenotype. To make parthenocarpic transgenic tomato plants without unwanted phenotypes, we found two genes, namely, Solyc03g007780 and Solyc02g067760, expressed in ovary tissue but not in vegetative tissues. Solyc03g007780 was expressed in developing ovaries and anthers. Solyc02g067760 mRNA was detected in whole-flower tissues. The promoters of Solyc03g007780 (Psol80) and Solyc02g067760 (Psol60) predominantly induced the expression of genes in the ovule, placenta, endocarp and pollen and in whole-flower tissues, respectively. Psol80/60-SlIAA9i lines, created for SlIAA9-RNA interference controlled by two promoters, successfully formed parthenocarpic fruits without pleiotropic effects in vegetative tissues. Downregulation of SlIAA9, responsible for parthenocarpic fruit formation, was observed in ovules rather than ovaries in the Psol80/60-SlIAA9i lines. Although the weight of parthenocarpic fruits of the Psol80/60-SlIAA9i lines was lower than the weight of pollinated fruits of the wild type (WT), the parthenocarpic fruits presented redder and more saturated colors and higher levels of total soluble solids and titratable acidity than the WT fruits.
| S-EPMC6906307 | biostudies-literature
Project description:The target of rapamycin complex (TORC) plays a key role in plant cell growth and survival by regulating the gene expression and metabolism according to environmental information. TORC activates transcription, mRNA translation, and anabolic processes under favorable conditions, thereby promoting plant growth and development. Tomato fruit ripening is a complex developmental process promoted by ethylene and specific transcription factors. TORC is known to modulate leaf senescence in tomato. In this study, we investigated the function of TORC in tomato fruit ripening using virus-induced gene silencing (VIGS) of the TORC genes, TOR, lethal with SEC13 protein 8 (LST8), and regulatory-associated protein of TOR (RAPTOR). Quantitative reverse transcription-polymerase chain reaction showed that the expression levels of tomato TORC genes were the highest in the orange stage during fruit development in Micro-Tom tomato. VIGS of these TORC genes using stage 2 tomato accelerated fruit ripening with premature orange/red coloring and decreased fruit growth, when control tobacco rattle virus 2 (TRV2)-myc fruits reached the mature green stage. TORC-deficient fruits showed early accumulation of carotenoid lycopene and reduced cellulose deposition in pericarp cell walls. The early ripening fruits had higher levels of transcripts related to fruit ripening transcription factors, ethylene biosynthesis, carotenoid synthesis, and cell wall modification. Finally, the early ripening phenotype in Micro-Tom tomato was reproduced in the commercial cultivar Moneymaker tomato by VIGS of the TORC genes. Collectively, these results demonstrate that TORC plays an important role in tomato fruit ripening by modulating the transcription of various ripening-related genes.
| S-EPMC9448650 | biostudies-literature
Project description:BACKGROUND: Fruit normally develops from the ovary after pollination and fertilization. However, the ovary can also generate seedless fruit without fertilization by parthenocarpy. Parthenocarpic fruit development has been obtained in tomato (Solanum lycopersicum) by genetic modification using auxin-synthesising gene(s) (DefH9-iaaM; DefH9-RI-iaaM) expressed specifically in the placenta and ovules. FINDINGS: We have performed a cDNA Amplified Fragment Length Polymorphism (cDNA-AFLP) analysis on pre-anthesis tomato flower buds (0.5 cm long) collected from DefH9-iaaM and DefH9-RI-iaaM parthenocarpic and wild-type plants, with the aim to identify genes involved in very early phases of tomato fruit development. We detected 212 transcripts differentially expressed in auxin-ipersynthesising pre-anthesis flower buds, 65 of them (31%) have unknown function. Several differentially expressed genes show homology to genes involved in protein trafficking and protein degradation via proteasome. These processes are crucial for auxin cellular transport and signaling, respectively. CONCLUSION: The data presented might contribute to elucidate the molecular basis of the fruiting process and to develop new methods to confer parthenocarpy to species of agronomic interest. In a recently published work, we have demonstrated that one of the genes identified in this screening, corresponding to #109 cDNA clone, regulates auxin-dependent fruit initiation and its suppression causes parthenocarpic fruit development in tomato.
| S-EPMC2718906 | biostudies-literature
Project description:Tomato fruits (Solanum lycopersicum L.) accumulate flavonoids in their cuticle and epidermal cells during ripening. These flavonoids come from de novo biosynthesis due to a significant increase in chalcone synthase (CHS) activity during ripening. Virus-induced gene silencing (VIGS) of tomato fruits have been used to down-regulate SlCHS expression during ripening and analyze the effects at the epidermal and cuticle level. Besides the expected change in fruit color due to a lack of flavonoids incorporated to the cuticle, several other modifications such as a decrease in the amount of cutin and polysaccharides were observed. These indicate a role for either flavonoids or CHS in the alteration of the expression levels of some genes involved in cuticle biosynthesis. Moreover, a negative interaction between the 2 cuticle components, flavonoids and waxes, suggests a relationship between these 2 metabolic pathways.
| S-EPMC4622847 | biostudies-other
Project description:BackgroundGeminiviruses encode few viral proteins. Most of the geminiviral proteins are multifunctional and influence various host cellular processes for the successful viral infection. Though few viral proteins like AC1 and AC2 are well characterized for their multiple functions, role of AC3 in the successful viral infection has not been investigated in detail.ResultsWe performed phage display analysis with the purified recombinant AC3 protein with Maltose Binding Protein as fusion tag (MBP-AC3). Putative AC3 interacting peptides identified through phage display were observed to be homologous to peptides of proteins from various metabolisms. We grouped these putative AC3 interacting peptides according to the known metabolic function of the homologous peptide containing proteins. In order to check if AC3 influences any of these particular metabolic pathways, we designed vectors for assaying DNA replication and virus induced gene-silencing of host gene PCNA. Investigation with these vectors indicated that AC3 enhances viral replication in the host plant tomato. In the PCNA gene-silencing experiment, we observed that the presence of functional AC3 ORF strongly manifested the stunted phenotype associated with the virus induced gene-silencing of PCNA in tomato plants.ConclusionsThrough the phage display analysis proteins from various metabolic pathways were identified as putative AC3 interacting proteins. By utilizing the vectors developed, we could analyze the role of AC3 in viral DNA replication and host gene-silencing. Our studies indicate that AC3 is also a multifunctional protein.
| S-EPMC3102638 | biostudies-literature