Harry Jonker

RNA Interference Silencing of Chalcone Synthase, the First Step in the Flavonoid Biosynthesis Pathway, Leads to Parthenocarpic Tomato Fruits

Parthenocarpy, the formation of seedless fruits in the absence of functional fertilization, is a desirable trait for several important crop plants, including tomato (Solanum lycopersicum). Seedless fruits can be of great value for consumers, the processing industry, and breeding companies. In this article, we propose a novel strategy to obtain parthenocarpic tomatoes by down-regulation of the flavonoid biosynthesis pathway using RNA interference (RNAi)-mediated suppression of chalcone synthase (CHS), the first gene in the flavonoid pathway. In CHS RNAi plants, total flavonoid levels, transcript levels of both Chs1 and Chs2, as well as CHS enzyme activity were reduced by up to a few percent of the corresponding wild-type values. Surprisingly, all strong Chs-silenced tomato lines developed parthenocarpic fruits. Although a relation between flavonoids and parthenocarpic fruit development has never been described, it is well known that flavonoids are essential for pollen development and pollen tube growth and, hence, play an essential role in plant reproduction. The observed parthenocarpic fruit development appeared to be pollination dependent, and Chs RNAi fruits displayed impaired pollen tube growth. Our results lead to novel insight in the mechanisms underlying parthenocarpic fruit development. The potential of this technology for applications in plant breeding and biotechnology will be discussed.

Production of Resveratrol in Recombinant Microorganisms

ABSTRACT Resveratrol production in Saccharomyces cerevisiae was compared to that in Escherichia coli. In both systems, 4-coumarate:coenzyme A ligase from tobacco and stilbene synthase from grapes were expressed. When p-coumaric acid was used as the precursor, resveratrol accumulations in the culture medium were observed to be comparable in E. coli (16 mg/liter) and yeast (6 mg/liter).

Purification and characterization of the enzymes of fructan biosynthesis in tubers of Helianthus tuberosus colombia. II. Purification of sucrose:sucrose 1-fructosyltransferase and reconstitution of fructan synthesis in vitro with purified sucrose:sucrose 1-fructosyltransferase and fructan-fructan 1-fructosyltransferase

Sucrose:sucrose 1-fructosyltransferase (1-SST), an enzyme involved in fructan biosynthesis, was purified to homogeneity from tubers of Helianthus tuberosus that were harvested in the accumulation phase. Gel filtration under native conditions predicted a molecular mass of about 67 kD. Electrophoresis or gel filtration under denaturing conditions yielded a 27- and a 55-kD fragment. 1-SST preferentially catalyzed the conversion of sucrose into the trisaccharide 1-kestose (GF2). Other reactions catalyzed by 1-SST at a lower rate were self-transfructosylations with GF2 and 1,1-nystose (GF3) as substrates yielding GF3 and 1,1,1-fructosylnystose, respectively, as products. 1-SST also catalyzed the removal of the terminal fructosyl unit from both GF2 and GF3, which resulted in the release of sucrose and GF2, respectively, and free Fru. The purified enzyme did not display [beta]-fructosidase activity. An enzyme mixture of purified 1-SST and fructan:fructan 1-fructosyltransferase, both isolated from tubers, was able to synthesize fructans up to a degree of polymerization of at least 13 with sucrose as a sole substrate.

Extensive metabolic cross-talk in melon fruit revealed by spatial and developmental combinatorial metabolomics

• Variations in tissue development and spatial composition have a major impact on the nutritional and organoleptic qualities of ripe fleshy fruit, including melon (Cucumis melo). To gain a deeper insight into the mechanisms involved in these changes, we identified key metabolites for rational food quality design. • The metabolome, volatiles and mineral elements were profiled employing an unprecedented range of complementary analytical technologies. Fruits were followed at a number of time points during the final ripening process and tissues were collected across the fruit flesh from rind to seed cavity. Approximately 2000 metabolite signatures and 15 mineral elements were determined in an assessment of temporal and spatial melon fruit development. • This study design enabled the identification of: coregulated hubs (including aspartic acid, 2-isopropylmalic acid, β-carotene, phytoene and dihydropseudoionone) in metabolic association networks; global patterns of coordinated compositional changes; and links of primary and secondary metabolism to key mineral and volatile fruit complements. • The results reveal the extent of metabolic interactions relevant to ripe fruit quality and thus have enabled the identification of essential candidate metabolites for the high-throughput screening of melon breeding populations for targeted breeding programmes aimed at nutrition and flavour improvement.

Metabolomic and elemental profiling of melon fruit quality as affected by genotype and environment

Melon (Cucumis melo L.) is a global crop in terms of economic importance and nutritional quality. The aim of this study was to explore the variability in metabolite and elemental composition of several commercial varieties of melon in various environmental conditions. Volatile and non-volatile metabolites as well as mineral elements were profiled in the flesh of mature fruit, employing a range of complementary analytical technologies. More than 1,000 metabolite signatures and 19 mineral elements were determined. Data analyses revealed variations related to factors such as variety, growing season, contrasting agricultural management practices (greenhouse vs. field with or without fruit thinning) and planting date. Two hundred and ninety-one analytes discriminated two contrasting varieties, one from the var. inodorous group and the other from the var. cantaloupensis group. Two hundred and eighty analytes discriminated a short shelf-life from a mid-shelf-life variety within the var. cantaloupensis group. Three hundred and twenty-seven analytes discriminated two seasons, and two hundred and fifty-two analytes discriminated two contrasting agricultural management practices. The affected compound families greatly depended on the factor studied. The compositional variability of identified or partially identified compounds was used to study metabolite and mineral element co-regulation using correlation networks. The results confirm that metabolome and mineral element profiling are useful diagnostic tools to characterize the quality of fruits cultivated under commercial conditions. They can also provide knowledge on fruit metabolism and the mechanisms of plant response to environmental modifications, thereby paving the way for metabolomics-guided improvement of cultural practices for better fruit quality.

Comprehensive metabolomics to evaluate the impact of industrial processing on the phytochemical composition of vegetable purees

The effects of conventional industrial processing steps on global phytochemical composition of broccoli, tomato and carrot purees were investigated by using a range of complementary targeted and untargeted metabolomics approaches including LC-PDA for vitamins, (1)H NMR for polar metabolites, accurate mass LC-QTOF MS for semi-polar metabolites, LC-MRM for oxylipins, and headspace GC-MS for volatile compounds. An initial exploratory experiment indicated that the order of blending and thermal treatments had the highest impact on the phytochemicals in the purees. This blending-heating order effect was investigated in more depth by performing alternate blending-heating sequences in triplicate on the same batches of broccoli, tomato and carrot. For each vegetable and particularly in broccoli, a large proportion of the metabolites detected in the purees was significantly influenced by the blending-heating order, amongst which were potential health-related phytochemicals and flavour compounds like vitamins C and E, carotenoids, flavonoids, glucosinolates and oxylipins. Our metabolomics data indicates that during processing the activity of a series of endogenous plant enzymes, such as lipoxygenases, peroxidases and glycosidases, including myrosinase in broccoli, is key to the final metabolite composition and related quality of the purees.

Multi-platform metabolomics analyses of a broad collection of fragrant and non-fragrant rice varieties reveals the high complexity of grain quality characteristics.

The quality of rice in terms not only of its nutritional value but also in terms of its aroma and flavour is becoming increasingly important in modern rice breeding where global targets are focused on both yield stability and grain quality. In the present paper we have exploited advanced, multi-platform metabolomics approaches to determine the biochemical differences in 31 rice varieties from a diverse range of genetic backgrounds and origin. All were grown under the specific local conditions for which they have been bred and all aspects of varietal identification and sample purity have been guaranteed by local experts from each country. Metabolomics analyses using 6 platforms have revealed the extent of biochemical differences (and similarities) between the chosen rice genotypes. Comparison of fragrant rice varieties showed a difference in the metabolic profiles of jasmine and basmati varieties. However with no consistent separation of the germplasm class. Storage of grains had a significant effect on the metabolome of both basmati and jasmine rice varieties but changes were different for the two rice types. This shows how metabolic changes may help prove a causal relationship with developing good quality in basmati rice or incurring quality loss in jasmine rice in aged grains. Such metabolomics approaches are leading to hypotheses on the potential links between grain quality attributes, biochemical composition and genotype in the context of breeding for improvement. With this knowledge we shall establish a stronger, evidence-based foundation upon which to build targeted strategies to support breeders in their quest for improved rice varieties.