Formate-tetrahydrofolate ligase: supplying the cytosolic one-carbon network in roots with one-carbon units originating from glycolate.

The metabolism of tetrahydrofolate (H4PteGlun)-bound one-carbon (C1) units (C1 metabolism) is multifaceted and required for plant growth, but it is unclear what of many possible synthesis pathways provide C1 units in specific organelles and tissues. One possible source of C1 units is via formate-tetrahydrofolate ligase, which catalyzes the reversible ATP-driven production of 10-formyltetrahydrofolate (10-formyl-H4PteGlun) from formate and tetrahydrofolate (H4PteGlun). Here, we report biochemical and functional characterization of the enzyme from Arabidopsis thaliana (AtFTHFL). We show that the recombinant AtFTHFL has lower Km and kcat values with pentaglutamyl tetrahydrofolate (H4PteGlu5) as compared to monoglutamyl tetrahydrofolate (H4PteGlu1), resulting in virtually identical catalytic efficiencies for the two substrates. Stable transformation of Arabidopsis plants with the EGFP-tagged AtFTHFL, followed with fluorescence microscopy, demonstrated cytosolic signal. Two independent T-DNA insertion lines with impaired AtFTHFL function had shorter roots compared to the wild type plants, demonstrating the importance of this enzyme for root growth. Overexpressing AtFTHFL led to the accumulation of H4PteGlun + 5,10-methylene-H4PteGlun and serine, accompanied with the depletion of formate and glycolate, in roots of the transgenic Arabidopsis plants. This metabolic adjustment supports the hypothesis that AtFTHFL feeds the cytosolic C1 network in roots with C1 units originating from glycolate, and that these units are then used mainly for biosynthesis of serine, and not as much for the biosynthesis of 5-methyl-H4PteGlun, methionine, and S-adenosylmethionine. This finding has implications for any future attempts to engineer one-carbon unit-requiring products through manipulation of the one-carbon metabolic network in non-photosynthetic organs.

Temperature differences between sites lead to altered phenylpropanoid metabolism in a varietal dependent manner.

Elevated temperature has already caused a significant loss of wine growing areas and resulted in inferior fruit quality, particularly in arid and semi-arid regions. The existence of broad genetic diversity in V. vinifera is key in adapting viticulture to climate change; however, a lack of understanding on the variability in berry metabolic response to climate change remains a major challenge to build ad-hoc strategies for quality fruit production. In the present study, we examined the impact of a consistent temperature difference between two vineyards on polyphenol metabolism in the berries of 20 red V. vinifera cultivars across three consecutive seasons (2017-2019). The results emphasize a varietal specific response in the content of several phenylpropanoid metabolites; the interaction factor between the variety and the vineyard location was also found significant. Higher seasonal temperatures were coupled with lower flavonol and anthocyanin contents, but such reductions were not related with the level of expression of phenylpropanoid related genes. Hierarchical clustering analyses of the metabolic data revealed varieties with a location specific response, exceptional among them was Tempranillo, suggesting a greater susceptibility to temperature of this cultivar. In conclusion, our results indicate that the extensive genetic capacity of V. vinifera bears a significant potential to withstand temperature increase associated with climate change.

The Effect of Topo-Climate Variation on the Secondary Metabolism of Berries in White Grapevine Varieties (Vitis vinifera).

Exploiting consistent differences in radiation and average air temperature between two experimental vineyards (Ramat Negev, RN and Mitzpe Ramon, MR), we examined the impact of climate variations on total carotenoids, redox status, and phenylpropanoid metabolism in the berries of 10 white wine grapevine (Vitis vinifera) cultivars across three consecutive seasons (2017-2019). The differences in carotenoid and phenylpropanoid contents between sites were seasonal and varietal dependent. However, the warmer RN site was generally associated with higher H2O2 levels and carotenoid degradation, and lower flavonol contents than the cooler MR site. Enhanced carotenoid degradation was positively correlated with radiation and daily degree days, leading to a greater drop in content from véraison to harvest in Colombard, Sauvignon Blanc, and Semillon berries. Analyses of berry H2O2 and phenylpropanoids suggested differences between cultivars in the links between H2O2 and flavonol contents. Generally, however, grapes with higher H2O2 content seem to have lower flavonol contents. Correlative network analyses revealed that phenylpropanoids at the warmer RN site are tightly linked to the radiation and temperature regimes during fruit ripening, indicating potentially harmful effect of warmer climates on berry quality. Specifically, flavan-3-ols were negatively correlated with radiation at RN. Principal component analysis showed that Muscat Blanc, Riesling, Semillon, and Sauvignon Blanc were the most site sensitive cultivars. Our results suggest that grapevine biodiversity is likely the key to withstand global warming hazards.

Metabolic Engineering Strategy Enables a Hundred-Fold Increase in Viniferin Levels in Vitis vinifera cv. Gamay Red Cell Culture.

Stilbenes are phytoalexins with health-promoting benefits for humans. Here, we boost stilbenes' production, and in particular the resveratrol dehydrodimer viniferin, with significant pharmacological properties, by overexpressing stilbene synthase (STS) under unlimited phenylalanine (Phe) supply. Vitis vinifera cell cultures were co-transformed with a feedback-insensitive E. coli DAHP synthase (AroG*) and STS genes, under constitutive promoters. All transgenic lines had increased levels of Phe and stilbenes (74-fold higher viniferin reaching 0.74 mg/g DW). External Phe feeding of AroG* + STS lines caused a synergistic effect on resveratrol and viniferin accumulation, achieving a 26-fold (1.33 mg/g DW) increase in resveratrol and a 620-fold increase (6.2 mg/g DW) in viniferin, which to date is the highest viniferin accumulation reported in plant cultures. We suggest that this strategy of combining higher Phe availability and STS expression generates grape cell cultures as potential factories for sustainable production of stilbenes with a minor effect on the levels of flavonoids.

Nitrogen availability and genotype affect major nutritional quality parameters of tef grain grown under irrigation.

Worldwide demand for tef (Eragrostis tef) as a functional food for human consumption is increasing, thanks to its nutritional benefits and gluten-free properties. As a result, tef in now grown outside its native environment in Ethiopia and thus information is required regarding plant nutrition demands in these areas, as well as resulting grain health-related composition. In the current work, two tef genotypes were grown in Israel under irrigation in two platforms, plots in the field and pots in a greenhouse, with four and five nitrogen treatments, respectively. Nutritional and health-related quality traits were analyzed, including mineral content, fatty acid composition, hydrophilic and lipophilic antioxidative capacity, total phenolic content and basic polyphenolic profile. Our results show that tef genotypes differ in their nutritional composition, e.g. higher phenolic contents in the brown compared to the white genotype. Additionally, nitrogen availability positively affected grain fatty acid composition and iron levels in both experiments, while negatively affecting total phenolics in the field trials. To conclude, nitrogen fertilization is crucial for crop growth and productivity, however it also implicates nutritional value of the grains as food. These effects should be considered when fertilizing tef with nitrogen, to optimize both crop productivity and nutritional effects.

Temperature Shift Between Vineyards Modulates Berry Phenology and Primary Metabolism in a Varietal Collection of Wine Grapevine.

Global climate change and the expected increase in temperature are altering the relationship between geography and grapevine (V. vinifera) varietal performance, and the implications of which are yet to be fully understood. We investigated berry phenology and biochemistry of 30 cultivars, 20 red and 10 white, across three seasons (2017-2019) in response to a consistent average temperature difference of 1.5°C during the growing season between two experimental sites. The experiments were conducted at Ramat Negev (RN) and Ramon (MR) vineyards, located in the Negev desert, Israel. A significant interaction between vineyard location, season, and variety affected phenology and berry indices. The warmer RN site was generally associated with an advanced phenological course for the white cultivars, which reached harvest up to 2 weeks earlier than at the MR site. The white cultivars also showed stronger correlation between non-consecutive phenological stages than did the red ones. In contrast, harvest time of red cultivars considerably varied according to seasons and sites. Warmer conditions extended fruit developmental phases, causing berry shriveling and cluster collapse in a few cultivars such as Pinot Noir, Ruby Cabernet, and Tempranillo. Analyses of organic acid content suggested differences between red and white cultivars in the content of malate, tartrate, and citrate in response to the temperature difference between sites. However, generally, cultivars at lower temperatures exhibited lower concentrations of pulp organic acids at véraison, but acid degradation until harvest was reduced, compared to the significant pace of acid decline at the warmer site. Sugars showed the greatest differences between sites in both white and red berries at véraison, but differences were seasonal dependent. At harvest, cultivars of both groups exhibited significant variation in hexose/sucrose ratio, and the averages of which varied from 1.6 to 2.9. Hexose/sucrose ratio was significantly higher among the red cultivars at the warmer RN, while this tendency was very slight among white cultivars. White cultivars seem to harbor a considerable degree of resilience due to a combination of earlier and shorter ripening phase, which avoids most of the summer heat. Taken together, our study demonstrates that the extensive genetic capacity of V. vinifera bears significant potential and plasticity to withstand the temperature increase associated with climate change.