SUMMARYMaturation of fleshy fruits such as tomato (Solanum lycopersicum) is subject to tight genetic control. Here we describe the development of a quantitative real-time PCR platform that allows accurate quantification of the expression level of approximately 1000 tomato transcription factors. In addition to utilizing this novel approach, we performed cDNA microarray analysis and metabolite profiling of primary and secondary metabolites using GC-MS and LC-MS, respectively. We applied these platforms to pericarp material harvested throughout fruit development, studying both wild-type Solanum lycopersicum cv. Ailsa Craig and the hp1 mutant. This mutant is functionally deficient in the tomato homologue of the negative regulator of the light signal transduction gene DDB1 from Arabidopsis, and is furthermore characterized by dramatically increased pigment and phenolic contents. We choose this particular mutant as it had previously been shown to have dramatic alterations in the content of several important fruit metabolites but relatively little impact on other ripening phenotypes. The combined dataset was mined in order to identify metabolites that were under the control of these transcription factors, and, where possible, the respective transcriptional regulation underlying this control. The results are discussed in terms of both programmed fruit ripening and development and the transcriptional and metabolic shifts that occur in parallel during these processes.
In seeds, glutamate decarboxylase (GAD) operates at the metabolic nexus between carbon and nitrogen metabolism by catalyzing the unidirectional decarboxylation of glutamate to form g-aminobutyric acid (GABA). To elucidate the regulatory role of GAD in seed development, we generated Arabidopsis (Arabidopsis thaliana) transgenic plants expressing a truncated GAD from Petunia hybrida missing the carboxyl-terminal regulatory Ca 2+ -calmodulin-binding domain under the transcriptional regulation of the seed maturation-specific phaseolin promoter. Dry seeds of the transgenic plants accumulated considerable amounts of GABA, and during desiccation the content of several amino acids increased, although not glutamate or proline. Dry transgenic seeds had higher protein content than wild-type seeds but lower amounts of the intermediates of glycolysis, glycerol and malate. The total fatty acid content of the transgenic seeds was 50% lower than in the wild type, while acyl-coenzyme A accumulated in the transgenic seeds. Labeling experiments revealed altered levels of respiration in the transgenic seeds, and fractionation studies indicated reduced incorporation of label in the sugar and lipid fractions extracted from transgenic seeds. Comparative transcript profiling of the dry seeds supported the metabolic data. Cellular processes up-regulated at the transcript level included the tricarboxylic acid cycle, fatty acid elongation, the shikimate pathway, tryptophan metabolism, nitrogen-carbon remobilization, and programmed cell death. Genes involved in the regulation of germination were similarly up-regulated. Taken together, these results indicate that the GAD-mediated conversion of glutamate to GABA during seed development plays an important role in balancing carbon and nitrogen metabolism and in storage reserve accumulation.Efficient assimilation of carbon (C) and nitrogen (N) is essential for optimal plant growth, productivity, and yield (Stitt, 1999), particularly in seeds in which the content of essential amino acids is low. Hence, a comprehensive understanding of seed development and metabolism is central to the enhancement of crop yield and quality. Seed development can be divided into three phases: cell division, maturation (accumulation of food reserves), and desiccation (Weber et al., 2005). The shift from one phase to the other is affected by sugars (Suc, hexoses, trehalose) and abscisic acid. In the early stages of seed development, maternal regulation is maintained via assimilate unloading and supply of nutrients, thereby sustaining cell division. During maturation, seed metabolism changes and storage reserves accumulate in expanding cells. At this time, photosynthetic activity is initiated in the seed, which is believed to improve oxygen supply and the energy state in the seed, thus counteracting increasingly hypoxic conditions (Borisjuk and Rolletschek, 2009). The maturation stage is followed by a phase termed "maturation drying," in which the metabolism of Arabidopsis (Arabidopsis thaliana) seeds shifts from a gen...
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