TERMINAL FLOWER-1/CENTRORADIALIS inhibits tuberisation via protein interaction with the tuberigen activation complex.

Potato tuber formation is a secondary developmental programme by which cells in the subapical stolon region divide and radially expand to further differentiate into starch-accumulating parenchyma. Although some details of the molecular pathway that signals tuberisation are known, important gaps in our knowledge persist. Here, the role of a member of the TERMINAL FLOWER 1/CENTRORADIALIS gene family (termed StCEN) in the negative control of tuberisation is demonstrated for what is thought to be the first time. It is shown that reduced expression of StCEN accelerates tuber formation whereas transgenic lines overexpressing this gene display delayed tuberisation and reduced tuber yield. Protein-protein interaction studies (yeast two-hybrid and bimolecular fluorescence complementation) demonstrate that StCEN binds components of the recently described tuberigen activation complex. Using transient transactivation assays, we show that the StSP6A tuberisation signal is an activation target of the tuberigen activation complex, and that co-expression of StCEN blocks activation of the StSP6A gene by StFD-Like-1. Transcriptomic analysis of transgenic lines misexpressing StCEN identifies early transcriptional events in tuber formation. These results demonstrate that StCEN suppresses tuberisation by directly antagonising the function of StSP6A in stolons, identifying StCEN as a breeding marker to improve tuber initiation and yield through the selection of genotypes with reduced StCEN expression.

Solutions to decrease a systematic error related to AAPH addition in the fluorescence-based ORAC assay.

Oxygen radical absorbance capacity (ORAC) assay in 96-well multi-detection plate readers is a rapid method to determine total antioxidant capacity (TAC) in biological samples. A disadvantage of this method is that the antioxidant inhibition reaction does not start in all of the 96 wells at the same time due to technical limitations when dispensing the free radical-generating azo initiator 2,2'-azobis (2-methyl-propanimidamide) dihydrochloride (AAPH). The time delay between wells yields a systematic error that causes statistically significant differences in TAC determination of antioxidant solutions depending on their plate position. We propose two alternative solutions to avoid this AAPH-dependent error in ORAC assays.

Increase in transcript accumulation of Psy1 and e-Lcy genes in grain development is associated with differences in seed carotenoid content between durum wheat and tritordeum.

Carotenoid rich diets have been associated with lower risk of certain diseases. The great importance of cereals in human diet has directed breeding programs towards carotenoid enhancement to alleviate these deficiencies in developing countries and to offer new functional foods in the developed ones. The new cereal tritordeum (×Tritordeum Ascherson et Graebener) derived from durum wheat (Triticum turgidum ssp. durum) and the wild barley Hordeum chilense, naturally presents carotenoid levels 5-8 times higher than those of durum wheat. The improvement of tritordeum properties as a new functional food requires the elucidation of biosynthetic steps for carotenoid accumulation in seeds that differ from durum wheat. In this work expression patterns of nine genes from the isoprenoid and carotenoid biosynthetic pathways were monitored during grain development in durum wheat and tritordeum. Additionally, a fine identification and quantification of pigments (chlorophylls and carotenoids) during grain development and in mature seeds has been addressed. Transcript levels of Psy1, Psy2, Zds, e-Lcy and b-Lcy were found to correlate to carotenoid content in mature grains. The specific activation of the homeologous genes Psy1, e-Lcy from H. chilense and the high lutein esterification found in tritordeum may serve to explain the differences with durum wheat in carotenoid accumulation.

Symbiotic fungi from a wild grass (Celtica gigantea) increase the growth, grain yield and quality of tritordeum under field conditions.

Plants function in symbiosis with numerous microorganisms, which might contribute to their adaptation and performance. In this study, we tested whether fungal strains in symbiotic interaction with roots of Celtica gigantea, a wild grass adapted to nutrient-poor soils in semiarid habitats, could improve the field performance of the agricultural cereal tritordeum (Triticum durum × Hordeum chilense). Seedlings of tritordeum were inoculated with 12 different fungal strains isolated from roots of Celtica gigantea that were first proved to promote the growth of tritordeum plants under greenhouse conditions. The inoculated seedlings were transplanted to field plots at two locations belonging to different climatic zones in terms of mean temperatures and precipitation in the Iberian Peninsula. Only one strain, Diaporthe iberica T6, had a significant effect on plant height, number of tillers and grain yield in one location. This result showed a substantial divergence between the results of greenhouse and field tests. In terms of grain nutritional quality, several parameters were differentially affected at both locations: Diaporthe T6, Pleosporales T7, Zygomycota T29 and Zygomycota T80 increased the content of total carotenoids, mainly lutein, in the colder location; whereas gluten proteins increased with several treatments in the warmer location. In conclusion, early inoculation of tritordeum plants with fungal symbionts had substantial beneficial effects on subsequent plant growth and development in the field. Regarding grain nutritional quality, the effect of inoculation was affected by the agroclimatic differences between both field locations.

Toward the Design of Potato Tolerant to Abiotic Stress.

Potato is a major global crop that has an important role to play in food security, reducing poverty and improving human nutrition. Productivity in potato however is limited in many environments by its sensitivity to abiotic stresses such as elevated temperature, drought, frost, and salinity. In this chapter we focus on the effects of elevated temperature on potato yields as high temperature is the most important uncontrollable factor affecting growth and yield of potato. We describe some of the physiological impacts of elevated temperature and review recent findings about response mechanisms. We describe genetic approaches that could be used to identify allelic variants of genes that may be useful to breed for increased climate resilience, an approach that could be deployed with recent advances in potato breeding.

Screening fungal endophytes from a wild grass for growth promotion in tritordeum, an agricultural cereal.

Celtica gigantea(= Stipa gigantea) is a large perennial grass which grows in nutrient-poor sandy soils in semiarid zones of the western Iberian Peninsula. The purpose of this work was to find out if culturable fungal symbionts isolated from roots of this wild grass could have growth promoting activity in tritordeum, a hybrid cereal for human consumption. A survey of fungi from the root endosphere of C. gigantea produced an isolate collection consisting of 60 different taxa, mostly ascomycetes. Fungal strains were inoculated into tritordeum plants in order to evaluate their effect in leaf and root biomass, nutrient content, and total antioxidant capacity. Two consecutive screening processes were made to test endophyte effects in plants. In the first screening, 66 strains were inoculated into seedlings by dipping roots in a liquid suspension of inoculum. In the second screening, 13 strains selected from the first screening were inoculated by sowing seeds in a substrate containing inoculum. The inoculation method used in the second screening involved less labor and plant manipulation and improved the quantity and quality of the inoculum, making it more appropriate for big scale experimental inoculation procedures. Several fungal strains promoted leaf or root growth. In particular, a strain belonging to the genus Diaporthe caused an increase in leaf and root biomass in both screening processes, suggesting that this endophyte might have a good potential for field application in tritordeum.

Effect of long-term storage on the free and esterified carotenoids in durum wheat (Triticum turgidum conv. durum) and tritordeum (×Tritordeum Ascherson et Graebner) grains.

The effect of long-term storage on the carotenoid composition in durum wheat and tritordeum grains was studied. Total carotenoid (mainly lutein) content decreased according to a temperature dependent first-order degradative kinetic model. The carotenoid retention was similar in both genotypes at the lower temperatures (71-73% at -32°C, 70% at 6°C and 55-56% at 20°C), whereas at the higher temperatures the pigment retention was higher in tritordeum (42% at 37°C; 10% at 50°C) than durum wheat (23% at 37°C; 1% at 50°C), probably due to the greater proportion of esterified xanthophylls. A clear difference between free and esterified pigments was observed, with smaller losses and slower degradation (higher stability) observed for the latter. The xanthophyll esterification process was highly specific and had a significant effect on the carotenoid stability according to the fatty acids involved in the esterification and their position on the lutein molecule. The results were consistent with a degradation process in which the carotenoid behavior is influenced by the chemical structure. Xanthophyll esterification can be promoted by environmental conditions, regardless of the cereal genotype, and is a powerful tool to modulate the carotenoid profile in cereals.

Lutein Esterification in Wheat Flour Increases the Carotenoid Retention and Is Induced by Storage Temperatures.

The present study aimed to evaluate the effects of long-term storage on the carotenoid pigments present in whole-grain flours prepared from durum wheat and tritordeum. As expected, higher storage temperatures showed a catabolic effect, which was very marked for free carotenoid pigments. Surprisingly, for both cereal genotypes, the thermal conditions favoured the synthesis of lutein esters, leading to an enhanced stability, slower degradation, and, subsequently, a greater carotenoid retention. The putative involvement of lipase enzymes in lutein esterification in flours is discussed, particularly regarding the preferential esterification of the hydroxyl group with linoleic acid at the 3' in the ε-ring of the lutein molecule. The negative effects of processing on carotenoid retention were less pronounced in durum wheat flours, which could be due to an increased esterifying activity (the de novo formation of diesterified xanthophylls was observed). Moreover, clear differences were observed for tritordeum depending on whether the lutein was in a free or esterified state. For instance, lutein-3'-O-monolinoleate showed a three-fold lower degradation rate than free lutein at 37 °C. In view of our results, we advise that the biofortification research aimed at increasing the carotenoid contents in cereals should be based on the selection of varieties with an enhanced content of esterified xanthophylls.

Carotenoid evolution during short-storage period of durum wheat (Triticum turgidum conv. durum) and tritordeum (×Tritordeum Ascherson et Graebner) whole-grain flours.

This study investigates the effect of storage temperature on carotenoid composition in durum wheat and tritordeum whole-grain flours. For both cereal genotypes, total carotenoid content significantly decreased during storage, following a temperature dependent first-order kinetic model. Individual and total carotenoid content decay were similar for durum wheat, with a maximum at 50 °C at the end of the storage period (94%). In contrast, the evolution of lutein ester fractions in tritordeum showed lower losses than for free lutein (∼ 50%), and consequently the total carotenoid content was less affected (83%). A decrease in the lutein monoesters fraction was observed, coinciding with an increase in the diesterified forms, especially for lutein dilinoleate. These data suggest an esterifying activity in flours different from the enzyme systems operating in vivo (xanthophyll acyl transferase). The formation of lutein diesters, with greater stability, explains the slower carotenoid degradation in tritordeum whole-grain flours.

Isolation and identification of lutein esters, including their regioisomers, in tritordeum (×Tritordeum Ascherson et Graebner) grains: Evidence for a preferential xanthophyll acyltransferase activity.

Liquid chromatography in conjunction with UV-visible spectroscopy and atmospheric pressure chemical ionisation (APCI) mass spectrometry has been used for the structural assignment of the lutein esters, including the regioisomeric forms, naturally occurring in the endosperm of tritordeum (×Tritordeum Ascherson et Graebner), a novel cereal. The distinctive mass spectrometry fragmentation pattern of lutein, characterized by a favored loss of the moieties at the position 3' of the ε-end ring, allowed an unambiguous structural identification of four monoesters (lutein 3'-O-linoleate, lutein 3-O-linoleate, lutein 3'-O-palmitate, lutein 3-O-palmitate) and four diesters (lutein dilinoleate, lutein 3'-O-linoleate-3-O-palmitate, lutein 3'-O-palmitate-3-O-linoleate, lutein dipalmitate). This is the first time that the regioisomers of carotenoid esters have been identified in a cereal. Evidences for a preferential xanthophyll acyltransferase activity regarding the position (3 or 3') and the acyl moiety are discussed. Further studies should be carried out in order to identify the acyltransferase enzymes and the acyl donor molecules involved in the xanthophyll esterification process.