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Analysis of subcellular metabolite levels of potato tubers (Solanum tuberosum) displaying alterations in cellular or extracellular sucrose metabolism.

ABSTRACT: The expression of a heterologous invertase in potato tubers (Solanum tuberosum) in either the cytosol or apoplast leads to a decrease in total sucrose content and to an increase in glucose. Depending on the targeting of the enzyme different changes in phenotype and metabolism of the tubers occur: the cytosolic invertase expressing tubers show an increase in the glycolytic flux, accumulation of amino acids and organic acids, and the appearance of novel disaccharides; however, these changes are not observed when the enzyme is expressed in the apoplast [Roessner et al. (2001). Plant Cell, 13, 11-29]. The analysis of these lines raised several questions concerning the regulation of compartmentation of metabolites in potato tubers. In the current study we addressed these questions by performing comparative subcellular metabolite profiling. We demonstrate that: (i) hexoses accumulate in the vacuole independently of their site of production, but that the cytosolic invertase expression led to a strong increase in the cytosolic glucose concentration and decrease in cytosolic sucrose, whereas these effects were more moderate in the apoplastic expressors; (ii) three out of four of the novel compounds found in the cytosolic overexpressors accumulate in the same compartment; (iii) despite changes in absolute cellular content the subcellular distribution of amino acids was invariant in the invertase overexpressing tubers. These results are discussed in the context of current models of the compartmentation of primary metabolism in heterotrophic plant tissues. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1007/s11306-008-0107-5) contains supplementary material, which is available to authorized users.

SUBMITTER: Farre EM

PROVIDER: S-EPMC2758360 | biostudies-literature | 2008 Jun

REPOSITORIES: biostudies-literature

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Analysis of subcellular metabolite levels of potato tubers (Solanum tuberosum) displaying alterations in cellular or extracellular sucrose metabolism.

Farre Eva M EM Fernie Alisdair R AR Willmitzer Lothar L

Metabolomics : Official journal of the Metabolomic Society 20080227 2

The expression of a heterologous invertase in potato tubers (Solanum tuberosum) in either the cytosol or apoplast leads to a decrease in total sucrose content and to an increase in glucose. Depending on the targeting of the enzyme different changes in phenotype and metabolism of the tubers occur: the cytosolic invertase expressing tubers show an increase in the glycolytic flux, accumulation of amino acids and organic acids, and the appearance of novel disaccharides; however, these changes are no ...[more]

PMID: 19816536

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Project description:BackgroundInvertases are ubiquitous enzymes that irreversibly cleave sucrose into fructose and glucose. Plant invertases play important roles in carbohydrate metabolism, plant development, and biotic and abiotic stress responses. In potato (Solanum tuberosum), invertases are involved in 'cold-induced sweetening' of tubers, an adaptive response to cold stress, which negatively affects the quality of potato chips and French fries. Linkage and association studies have identified quantitative trait loci (QTL) for tuber sugar content and chip quality that colocalize with three independent potato invertase loci, which together encode five invertase genes. The role of natural allelic variation of these genes in controlling the variation of tuber sugar content in different genotypes is unknown.ResultsFor functional studies on natural variants of five potato invertase genes we cloned and sequenced 193 full-length cDNAs from six heterozygous individuals (three tetraploid and three diploid). Eleven, thirteen, ten, twelve and nine different cDNA alleles were obtained for the genes Pain-1, InvGE, InvGF, InvCD141 and InvCD111, respectively. Allelic cDNA sequences differed from each other by 4 to 9%, and most were genotype specific. Additional variation was identified by single nucleotide polymorphism (SNP) analysis in an association-mapping population of 219 tetraploid individuals. Haplotype modeling revealed two to three major haplotypes besides a larger number of minor frequency haplotypes. cDNA alleles associated with chip quality, tuber starch content and starch yield were identified.ConclusionsVery high natural allelic variation was uncovered in a set of five potato invertase genes. This variability is a consequence of the cultivated potato's reproductive biology. Some of the structural variation found might underlie functional variation that influences important agronomic traits such as tuber sugar content. The associations found between specific invertase alleles and chip quality, tuber starch content and starch yield will facilitate the selection of superior potato genotypes in breeding programs.

Project description:1. Methylation analysis of potato (Solanum tuberosum) lectin and thorn-apple (Datura stramonium) lectin confirmed previous conclusions that both glycoproteins contained high proportions of l-arabinofuranosides and lesser amounts of d-galactopyranosides. The arabinofuranosides are present in both lectins as short unbranched chains containing 1-->2- and 1-->3-linkages, which are known to be linked to hydroxyproline. Galactopyranosides are present as monosaccharides, which are known to be attached to serine, in potato lectin and as both the monosaccharide and the 1-->3-linked disaccharide in Datura lectin. 2. Alkaline digestion of potato lectin and subsequent separation of the components by gel filtration led to the isolation of four fractions corresponding to the mono-, di-, tri- and tetra-arabinosides of hydroxyproline. The latter two fractions accounted for over 70% of the total hydroxyproline. 3. Methylation analysis was used to show that the triarabinoside contained only 1-->2-linkages between sugars, but that the tetra-arabinoside contained both 1-->2- and 1-->3-linkages. Direct-insertion mass spectrometry of these compounds using electron impact and chemical ionization, in a comparison with other known structural patterns, was used to determine the sequences of the sugars, which were Araf1-->2Araf1-->2Araf1-->Hyp and Araf1-->3Araf1-->2Araf1-->2Araf 1-->Hyp. 4. On the basis of optical rotation it had previously been suggested [Allen, Desai, Neuberger & Creeth (1978) Biochem. J.171, 665-674] that all the arabinose of potato lectin was present as the beta-l-furanoside. However, measurement of the optical rotations of the hydroxyprolyl arabinosides showed that whereas the diarabinoside had a molar rotation ([m]) value close to that predicted, the triarabinoside was more dextrorotatory and the tetra-arabinoside was less dextrorotatory than expected. Possible explanations for these findings are that, although the di- and tri-arabinosides contain exclusively beta-arabinofuranosides, in the tri-arabinoside, interactions between pentose units lead to an enhanced positive rotation. The tetra-arabinoside, however, is proposed to contain a single alpha-arabinofuranoside residue, which is responsible for the lower than expected positive rotation. The observed rotation of the tetra-arabinoside was found to be close to the theoretical value predicted on that basis. Furthermore, the action of a specific alpha-arabinofuranosidase on the tetrasaccharide was to remove a single arabinose residue, presumably the terminal non-reducing sugar, and to produce a product that was indistinguishable on electrophoresis from the triarabinoside. Changes in rotation were compatible with this assumption. 5. It is concluded that the structures of the hydroxyprolyl tri- and tetra-arabinosides of potato lectin are: betaAraf1-->2betaAraf1-->2betaAraf1-->Hyp and alphaAraf1-->3betaAraf1-->2betaAraf 1-->2betaAraf1-->Hyp. These are identical with compounds that have been isolated from the insoluble hydroxyproline-rich glycoproteins of plant cell walls.

Project description:Development of an efficient and eco-friendly technique to break tuber dormancy in potato (Solanum tuberosum L.) is highly demanded due to the production of two or more crops annually. Several physiological and hormonal changes have been found to be related to the breaking of tuber dormancy; however, their consistency with genotypes and different protocols have not been well clarified. This study aims to evaluate the effectiveness of four dormancy-breaking methods, that is, plant growth regulator (PGR) dipping in 30, 60, or 90 mgL-1 benzyl amino purine (BAP) and 10, 20, or 30 mgL-1 gibberellic acids (GA3) alone and in the combination of optimized concentrations; electric current application at 20, 40, 60, or 80 Vs; cold pre-treatment at 2, 4, or 6 °C; irradiation at 1, 1.5, 2, 2.5, 3, or 3.5 kGy. In addition, changes in endogenous levels of abscisic acid (ABA), zeatin (ZT), and gibberellin A1 (GA1) in six potato genotypes after subjecting to these methods were investigated. Overall, the highest effective method for dormancy duration was the PGR application which shortened the duration by 18 days, followed by electric current (13 days), cold pre-treatment (9 days), and then irradiation (7 days). The solution of 60 mgL-1 BAP significantly reduced the dormancy duration in all genotypes but did not have a significant effect on the sprout length. While 20 mgL-1 GA3 produced maximum sprout length with a non-significant effect on dormancy duration. The genotype × PGR interaction for dormancy duration was more pronounced in short- and medium-term dormancy genotypes than in long-term dormancy genotypes. The genotypes displayed a significant positive correlation between dormancy duration and ABA levels but exhibited a negative correlation between dormancy duration and ZT as well as GA1 levels. From the first to the third week of storage, ABA was decreased in tubers while, however, ZT and GA1 were increased. The obtained results could be useful for the postharvest storage of potato tuber and the related field of physiological investigation in future.

Dataset Information

Analysis of subcellular metabolite levels of potato tubers (Solanum tuberosum) displaying alterations in cellular or extracellular sucrose metabolism.

Publications

Analysis of subcellular metabolite levels of potato tubers (Solanum tuberosum) displaying alterations in cellular or extracellular sucrose metabolism.

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