Unknown,Transcriptomics,Genomics,Proteomics

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Molecular analysis of a tomato OGDH antisense line

ABSTRACT: Transgenic tomato (Solanum lycopersicum) plants expressing a fragment of the SlSIOGDH gene encoding subunit E1 of the 2-oxoglutarate dehydrogenase protein complex in the antisense orientation and exhibiting considerable reductions in the activity of this enzyme exhibit a considerably reduced rate of respiration. They were, however, characterised by largely unaltered photosynthetic rates and fruit yield but a restricted leaf, stem and root growth rate. The lines displayed markedly altered metabolic profiles including changes in tricarboxylic acid cycle intermediates and in the majority of the amino acids without alterations in pyridine nucleotide content. Moreover, they displayed a generally accelerated development exhibiting early flowering, accelerated fruit ripening and a markedly earlier onset of leaf senescence. In addition, transcript and selective hormone profiling of gibberellins and ABA revealed changes only in the former coupled to changes in transcripts encoding enzymes of gibberellins biosynthesis. The data obtained are discussed in the context of the importance of this enzyme in both photosynthetic and respiratory metabolism as well as in programs of plant development connected to carbon nitrogen interactions. Three biological replicates per genotype were analysed. Genotypes analyzed were tomato var. M82 wild type plants grown under green house conditions with one 2-oxo-glutarate dehydrogenase antisense line grown under the same conditions.

ORGANISM(S): Solanum lycopersicum

SUBMITTER: Marc Lohse

PROVIDER: E-GEOD-35618 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:Colletotrichum coccodes secretes ammonium during germination and colonization of host tissue as a mechanism for dual activation of fungal pathogenicity factors and plant cell death via activation of host NADPH oxidase. Ammonium accumulation during colonization of tomato fruits by C. coccodes induced host nucleus integrity changes and program cell death (PCD). Transcriptome analysis at the leading edge of colonized tissue and ammonium treated tomato fruits revealed 82 and 237 common up-regulated and common down-regulated genes, respectively. The common-up-regulated genes showed over representation of pathogen related (PR) proteins, salicylic acid (SA) dependent and systemic acquired resistance (SAR) related genes and genes related to biotic stress. The down regulated genes showed over representation of Jasmonic acid (JA) dependent genes. Consistent with this observation the direct application of SA enhanced C. coccodes colonization, whereas the application of JA delayed colonization. Candidate genes were selected for QRT-PCR analysis of transcripts in normal and transgenic RBOH antisense plants and showed dependence on RBOH regulation. In all, the results suggest that ammonia accumulation during C. coccodes colonization activates SA and suppress JA pathways through activation of RBOH. The effects of ammonium on the expression of the selected up-and down regulated genes were examined in fruit of different maturation stages. Red, susceptible and green resistant, tomato fruit displayed similar behavior, suggesting that resistance of green fruits to C. coccodes colonization is not modulated by the JA and SA pathways but depends on other factors Ten micrograms of total RNA was processed for the microarray hybridizations using the Affymetrix GeneChip one-cycle target-labeling kit (Affymetrix). The four treatments were: C. coccodes "mat inoculation" compared to a wound control (24 hours after the tomato fruit peel was removed). 5 mM Ammonium treatment in PBS compared to PBS treatments as a control for ammonium treatment, the treatments applied every four hours. Each treatment had a duplicate. The biotinylated complementary RNA was fragmented and hybridized to the GeneChip Tomato Genome Array (10,038 tomato probe sets for more than 9,200 tomato genes). The arrays were washed, stained, and scanned at the biological services (Weizmann institute of science, Rehovot, Israel). CEL files for the Affymetrix microarray data arising from the microarray experiments were analyzed utilizing the Partek genomics software (Downey, 2006).

Project description:Tomato fruit ripening is under the control of ethylene as well as a group of ethylene-independent transcription factors, including NON-RIPENING (NOR) and RIPENING INHIBITOR (RIN). During ripening, the linear carotene lycopene accumulates at the expense of cyclic carotenoids. Fruit-specific overexpression of LYCOPENE β-CYCLASE (LCYb) under the control of the PHYTOENE DESATURASE (PDS) promoter resulted in increased levels of β-carotene and ABA and in decreased ethylene levels. Genes regulated by ABA, or involved in its synthesis and signaling, were overexpressed, while those associated with ethylene and cell wall remodeling were repressed. In agreement with the transcriptional data, LCYb-overexpressing fruits exhibited increased density of cell wall material containing linear, under-methylated pectins and displayed an array of additional ripening phenotypes, including delayed softening, increased turgor, enhanced shelf life and a thicker cuticle with a higher content of cutin monomers and triterpenoids. The levels of several primary metabolites and phenylpropanoids also changed in the transgenics, which could be attributed to delayed fruit ripening and to ABA respectively. Network correlation analysis suggests that ABA, acting through NOR and RIN, is responsible for many of the above phenotypes. These data reinforce suggestions that ABA plays an important role in tomato fruit ripening and provide clues that fruit b-carotene, acting as a precursor for ABA, actively participates in controlling the ripening process rather than merely being an output thereof. Overexpression of a LCYb gene from Arabidopsis under the control of the ripening-associated PDS promoter leads to ripe tomato fruits accumulating high β-carotene levels. Using several independent transgenic lines, we conducted a system-wide study of the effect of increased β-carotene levels on tomato fruit ripening and shelf life. Our data suggest that β-carotene, acting through ABA, is involved in a regulatory loop within the network controlling tomato fruit ripening.

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Molecular analysis of a tomato OGDH antisense line

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