Integrative Transcript and Metabolite Analysis of Nutritionally Enhanced DE-ETIOLATED1 Downregulated Tomato Fruit

Centre for Systems and Synthetic Biology, University of London, Egham, Surrey TW20 0EX, United Kingdom.
The Plant Cell (Impact Factor: 9.34). 04/2010; 22(4):1190-215. DOI: 10.1105/tpc.110.073866
Source: PubMed


Fruit-specific downregulation of the DE-ETIOLATED1 (DET1) gene product results in tomato fruits (Solanum lycopersicum) containing enhanced nutritional antioxidants, with no detrimental effects on yield. In an attempt to further our understanding of how modulation of this gene leads to improved quality traits, detailed targeted and multilevel omic characterization has been performed. Metabolite profiling revealed quantitative increases in carotenoid, tocopherol, phenylpropanoids, flavonoids, and anthocyanidins. Qualitative differences could also be identified within the phenolics, including unique formation in fruit pericarp tissues. These changes resulted in increased total antioxidant content both in the polar and nonpolar fractions. Increased transcription of key biosynthetic genes is a likely mechanism producing elevated phenolic-based metabolites. By contrast, high levels of isoprenoids do not appear to result from transcriptional regulation but are more likely related to plastid-based parameters, such as increased plastid volume per cell. Parallel metabolomic and transcriptomic analyses reveal the widespread effects of DET1 downregulation on diverse sectors of metabolism and sites of synthesis. Correlation analysis of transcripts and metabolites independently indicated strong coresponses within and between related pathways/processes. Interestingly, despite the fact that secondary metabolites were the most severely affected in ripe tomato fruit, our integrative analyses suggest that the coordinated activation of core metabolic processes in cell types amenable to plastid biogenesis is the main effect of DET1 loss of function.

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    • "Research quantitative increases in carotenoids, tocopherol, phenylpropanoids , flavonoids and anthocyanins which were positively correlated with increased transcription of key biosynthetic genes of these metabolic processes (Enfissi et al., 2010). Transcription factor profiling and metabolite profiling, combined with microarray analysis, were also performed using different stages of hp1 fruit, which revealed that the secondary metabolism changes were consistent with the expression dynamics of transcription factors (Rohrmann et al., 2011). "
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    ABSTRACT: In tomato (Solanum lycopersicum), high pigment mutations (hp-1 and hp-2) were mapped to genes encoding UV-damaged DNA binding protein 1 (DDB1) and de-etiolated-1 (DET1), respectively. Here we characterized a tomato methyl-CpG-binding domain protein SlMBD5 identified by yeast two-hybrid screening using SlDDB1 as a bait. Yeast two-hybrid assay demonstrated that the physical interaction of SlMBD5 with SlDDB1 is mediated by the C-termini of SlMBD5 and the β-propeller-C (BPC) of SlDDB1. Co-immunoprecipitation analyses revealed that SlMBD5 associates with SlDDB1-interacting partners including SlDET1, SlCUL4, SlRBX1a and SlRBX1b in vivo. SlMBD5 was shown to target to nucleus and dimerizes via its MBD motif. Electrophoresis mobility shift analysis suggested that the MBD of SlMBD5 specifically binds to methylated CpG dinucleotides but not to methylated CpHpG or CpHpH dinucleotides. SlMBD5 expressed in protoplast is capable of activating transcription of CG islands, whereas CUL4/DDB1 antagonizes this effect. Overexpressing SlMBD5 resulted in diverse developmental alterations including darker green fruits with increased plastid level and elevated pigmentation, as well as enhanced expression of SlGLK2, a key regulator of plastid biogenesis. Taken together, we hypothesize that the physical interaction of SlMBD5 with the CUL4-DDB1-DET1 complex component may affect its binding activity to methylated DNA and subsequently attenuate its transcription activation of downstream genes.
    Full-text · Article · Nov 2015 · New Phytologist
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    • "SlCUL4, SlDDB1 and SlDET1 have been shown previously to function as negative regulators of plastid level and pigmentation in tomato (Mustilli et al., 1999; Lieberman et al., 2004; Liu et al., 2004; Wang et al., 2008; Enfissi et al., 2010; Powell et al., 2012; Nguyen et al., 2014). This is antagonistic to the role of the positive regulator, tomato SlGLK2 that promotes chloroplast development and metabolite accumulation (Powell et al., 2012; Nguyen et al., 2014). "
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    ABSTRACT: CULLIN4-RING ubiquitin ligases (CRL4s) as well as their targets are fundamental regulators functioning in many key developmental and stress responses in eukaryotes. In tomato (Solanum lycopersicum), molecular cloning has revealed that the underlying genes of natural spontaneous mutations high pigment 1 (hp1), high pigment 2 (hp2) and uniform ripening (u) encode UV-DAMAGED DNA BINDING PROTEIN 1 (DDB1), DE-ETIOLATED 1 (DET1) and GOLDEN 2-LIKE (GLK2), respectively. However, the molecular basis of the opposite actions of tomato GLK2 vs CUL4-DDB1-DET1 complex on regulating plastid level and fruit quality remains unknown. Here, we provide molecular evidence showing that the tomato GLK2 protein is a substrate of the CUL4-DDB1-DET1 ubiquitin ligase complex for the proteasome degradation. SlGLK2 is degraded by the ubiquitin-proteasome system, which is mainly determined by two lysine residues (K11 and K253). SlGLK2 associates with the CUL4-DDB1-DET1 E3 complex in plant cells. Genetically impairing CUL4, DDB1 or DET1 results in a retardation of SlGLK2 degradation by the 26S proteasome. These findings are relevant to the potential of nutrient accumulation in tomato fruit by mediating the plastid level and contribute to a deeper understanding of an important regulatory loop, linking protein turnover to gene regulation.
    Full-text · Article · Sep 2015 · New Phytologist
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    • "It was documented that photomorphogenesis may be indirectly associated with the flavonoid pathway. Suppression of the DET1 gene led to increased carotenoid and flavonoid levels in tomato fruits, as well as phenylpropanoid and anthocyanidins (Davuluri et al., 2005; Enfissi et al., 2010). It is well known that HY5 plays an important role in photomorphogenic development (Chattopadhyay et al., 1998). "

    Full-text · Article · Sep 2015 · Frontiers in Plant Science
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