Raquel Olías

The plasma membrane Na+/H+ antiporter SOS1 is essential for salt tolerance in tomato and affects the partitioning of Na+ between plant organs

We have identified a plasma membrane Na(+)/H(+) antiporter gene from tomato (Solanum lycopersicum), SlSOS1, and used heterologous expression in yeast to confirm that SlSOS1 was the functional homolog of AtSOS1. Using post-transcriptional gene silencing, we evaluated the role played by SlSOS1 in long-distance Na(+) transport and salt tolerance of tomato. Tomato was used because of its anatomical structure, more complex than that of Arabidopsis, and its agricultural significance. Transgenic tomato plants with reduced expression of SlSOS1 exhibited reduced growth rate compared to wild-type (WT) plants in saline conditions. This sensitivity correlated with higher accumulation of Na(+) in leaves and roots, but lower contents in stems of silenced plants under salt stress. Differential distribution of Na(+) and lower net Na(+) flux were observed in the xylem sap in the suppressed plants. In addition, K(+) concentration was lower in roots of silenced plants than in WT. Our results demonstrate that SlSOS1 antiporter is not only essential in maintaining ion homeostasis under salinity, but also critical for the partitioning of Na(+) between plant organs. The ability of tomato plants to retain Na(+) in the stems, thus preventing Na(+) from reaching the photosynthetic tissues, is largely dependent on the function of SlSOS1.

Two closely linked tomato HKT coding genes are positional candidates for the major tomato QTL involved in Na+/K+ homeostasis

The location of major quantitative trait loci (QTL) contributing to stem and leaf [Na(+) ] and [K(+) ] was previously reported in chromosome 7 using two connected populations of recombinant inbred lines (RILs) of tomato. HKT1;1 and HKT1;2, two tomato Na(+) -selective class I-HKT transporters, were found to be closely linked, where the maximum logarithm of odds (LOD) score for these QTLs located. When a chromosome 7 linkage map based on 278 single-nucleotide polymorphisms (SNPs) was used, the maximum LOD score position was only 35 kb from HKT1;1 and HKT1;2. Their expression patterns and phenotypic effects were further investigated in two near-isogenic lines (NILs): 157-14 (double homozygote for the cheesmaniae alleles) and 157-17 (double homozygote for the lycopersicum alleles). The expression pattern for the HKT1;1 and HKT1;2 alleles was complex, possibly because of differences in their promoter sequences. High salinity had very little effect on root dry and fresh weight and consequently on the plant dry weight of NIL 157-14 in comparison with 157-17. A significant difference between NILs was also found for [K(+) ] and the [Na(+) ]/[K(+) ] ratio in leaf and stem but not for [Na(+) ] arising a disagreement with the corresponding RIL population. Their association with leaf [Na(+) ] and salt tolerance in tomato is also discussed.

Overexpression of SlSOS2 (SlCIPK24) confers salt tolerance to transgenic tomato

The Ca(2+)-dependent SOS pathway has emerged as a key mechanism in the homeostasis of Na(+) and K(+) under saline conditions. We have identified and functionally characterized the gene encoding the calcineurin-interacting protein kinase of the SOS pathway in tomato, SlSOS2. On the basis of protein sequence similarity and complementation studies in yeast and Arabidopsis, it can be concluded that SlSOS2 is the functional tomato homolog of Arabidopsis AtSOS2 and that SlSOS2 operates in a tomato SOS signal transduction pathway. The biotechnological potential of SlSOS2 to provide salt tolerance was evaluated by gene overexpression in tomato (Solanum lycopersicum L. cv. MicroTom). The better salt tolerance of transgenic plants relative to non-transformed tomato was shown by their faster relative growth rate, earlier flowering and higher fruit production when grown with NaCl. The increased salinity tolerance of SlSOS2-overexpressing plants was associated with higher sodium content in stems and leaves and with the induction and up-regulation of the plasma membrane Na(+)/H(+) (SlSOS1) and endosomal-vacuolar K(+), Na(+)/H(+) (LeNHX2 and LeNHX4) antiporters, responsible for Na(+) extrusion out of the root, active loading of Na(+) into the xylem, and Na(+) and K(+) compartmentalization.

The Na+/H+ exchanger SOS1 controls extrusion and distribution of Na+ in tomato plants under salinity conditions

Maintaining a high K+/Na+ ratio in the cell cytosol, along with the transport processes implicated in the xylem and phloem loading/unloading of Na+ in plants (long-distance transport) are key aspects in plant salt tolerance. The Ca2+-dependent SOS pathway regulating Na+ and K+ homeostasis and long-distance Na+ transport has been reported in Arabidopsis. However, Arabidopsis might not be the best model to analyze the involvement of the SOS pathway in long-distance Na+ transport due to the very short stem of these plants which do not allow a precise dissection of the relative content of Na+ in stem versus leaf. This separation would be critical to assess the role of SOS1 in xylem loading/unloading, Na+ export by roots, retention in stems and the differential distribution/accumulation in old leaves. Towards this goal, tomato might represent a superior model due to its anatomical structure and agricultural significance. We recently demonstrated the key role played by the plasma membrane Na+/H+ antiporter SlSOS1 in salt tolerance in tomato by maintaining ion homeostasis under salinity stress and in the partitioning of Na+ in plant organs.

Modified atmosphere packaging of strawberry fruit: Effect of package perforation on oxygen and carbon dioxide / Envasado de fresas en atmósfera modificada: Efecto de la perforación del envase en el contenido de oxígeno y dióxido de carbono:

The effect of film perforation on gas content during modified atmosphere packaging of strawberry fruit was studied. Camarosa strawberries were refrigerated at 2 °C, and wrapped with polypropy lene (PP) film with perforations of 1 and 2 mm diameter. Fruits were held at 2 °C during three days, simulating refrigerated transport, and then at 20 °C for four days to simulate retail display tempera tures. Perforation degree affected final gas contents inside the packages, ranging from 5.68-25.96% CO2 and from 4.84-15.69% O2 on day 7. Different gas content profiles were found among packages with similar perforated surface but distinct diameter perforations, which could infer a perforation edge effect on gas exchange. Neither a CO, stratification effect nor a gas exchange site effect were observed for this kind of perforation-mediated modified atmosphere packaging of strawberries.

Strawberry quality as a function of the ‘high pressure fast cooling’ design

Abstract Oso Grande strawberries were picked, wrapped with polyvinyl chloride film, and finally cooled by ‘high pressure fast cooling’ units with slight differences in design in terms of number of fans functioning and ventilation spacer widths. The effect on strawberry quality of these cooling cells was evaluated compared to room-cooled fruits, and when compared to quality parameter values at fruit gathering. Quality evaluations by means of determination of main sugars and organic acids, colour, firmness, total soluble solids, titrable acidity and pH assessments were carried out at different commercial stages, immediately after cooling, after 3 days at 2 °C simulating refrigerated transport, and after 4 days at 17 °C storage (shelf-life). Reducing (by half) the ventilation spacer (V cell), resulted in a 20% higher air speed and a 20% faster cooling than control units (N cell), while units using half of the fans (F cell) showed a lower air speed (45%) and a 30% more time to cool down the product. These modifications also affected strawberry quality. Fruits from the fastest cooler cell, V cell, showed a different quality pattern compared to fruits from N and F cells, with a higher sucrose inversion and less glucose and fructose contents. V cell-cooled strawberries presented a lower content of citric, malic and ascorbic acids during shelf-life. However, fruits cooled by the V cell showed the highest marketable fruit percentage, although with a decrease in red colour compared to fruit at gathering, and fruit cooled by the N or F cells. In these cells, fruit colour remained practically unchanged along the different commercial stages including the long shelf-life period.

Eliminating Anti-Nutritional Plant Food Proteins: The Case of Seed Protease Inhibitors in Pea

Several classes of seed proteins limit the utilisation of plant proteins in human and farm animal diets, while plant foods have much to offer to the sustainable intensification of food/feed production and to human health. Reduction or removal of these proteins could greatly enhance seed protein quality and various strategies have been used to try to achieve this with limited success. We investigated whether seed protease inhibitor mutations could be exploited to enhance seed quality, availing of induced mutant and natural Pisum germplasm collections to identify mutants, whilst acquiring an understanding of the impact of mutations on activity. A mutant (TILLING) resource developed in Pisum sativum L. (pea) and a large germplasm collection representing Pisum diversity were investigated as sources of mutations that reduce or abolish the activity of the major protease inhibitor (Bowman-Birk) class of seed protein. Of three missense mutations, predicted to affect activity of the mature trypsin / chymotrypsin inhibitor TI1 protein, a C77Y substitution in the mature mutant inhibitor abolished inhibitor activity, consistent with an absolute requirement for the disulphide bond C77-C92 for function in the native inhibitor. Two further classes of mutation (S85F, E109K) resulted in less dramatic changes to isoform or overall inhibitory activity. The alternative strategy to reduce anti-nutrients, by targeted screening of Pisum germplasm, successfully identified a single accession (Pisum elatius) as a double null mutant for the two closely linked genes encoding the TI1 and TI2 seed protease inhibitors. The P. elatius mutant has extremely low seed protease inhibitory activity and introgression of the mutation into cultivated germplasm has been achieved. The study provides new insights into structure-function relationships for protease inhibitors which impact on pea seed quality. The induced and natural germplasm variants identified provide immediate potential for either halving or abolishing the corresponding inhibitory activity, along with associated molecular markers for breeding programmes. The potential for making large changes to plant protein profiles for improved and sustainable food production through diversity is illustrated. The strategy employed here to reduce anti-nutritional proteins in seeds may be extended to allergens and other seed proteins with negative nutritional effects. Additionally, the novel variants described for pea will assist future studies of the biological role and health-related properties of so-called anti-nutrients.

Involvement of SlSOS2 in tomato salt tolerance.

The Ca2+-dependent SOS pathway has emerged as a key mechanism in the homeostasis of Na+ and K+ under saline conditions. We recently identified and functionally characterized by complementation studies in yeast and Arabidopsis the gene encoding the calcineurin-interacting protein kinase of the SOS pathway in tomato, SlSOS2.1 We also show evidences on the biotechnological potential of SlSOS2 conferring salt tolerance to transgenic tomato. The increased salinity tolerance of SlSOS2 overexpressing plants is associated with higher sodium content in stems and leaves. SlSOS2 overexpression upregulates the Na+/H+ exchange at the plasma membrane (SlSOS1) and K+,Na+/H+ antiport at the endosomal and vacuolar compartments (LeNHX2 and LeNHX4). Therefore, SlSOS2 seems to be involved in tomato salinity tolerance through regulation of Na+ extrusion from the root, active loading of Na+ into the xylem and Na+ and K+ compartmentalization.

Quality Assessment of Strawberries Packed with Perforated Polypropylene Punnets During Cold Storage

Strawberries stored in polypropylene packages with different perforation surfaces were maintained for three weeks at 2°C simulating long refrigerated transport or long cold storage. After 3 days at 2°C, CO2 levels inside the packages were in the range of 1.5—2.6%, while O2 contents were 17.8—18.9% for the three perforation surfaces under study. Gas levels remained practically unchanged for one week and then changed slowly such that at day 21 they were 7.2—8.8% and 13.6—14.9% for CO2 and O2, respectively. There was a suitable fruit quality maintenance during the first 10 days at 2°C. After this period of time there was an increased risk of fruit quality impairment, with an increasing marketable fruit loss and off-flavors development, and a slight decrease of sugar and organic acids content. There were practically no differences among the three microperforated packages studied in terms of fruit quality during this recommended 10-day period at 2°C with low levels of marketable fruit loss, no signal of Botrytis infection, and only a slight decrease in sucrose content. Comparison with control fruits during the first 10 days at 2°C showed practically no differences except for this microperforated packaging system being associated to lower anthocyanin content in the fruits. Se almacenaron fresas en envases de propileno perforados con diferente tamaño de orificios durante 3 semanas a 2°C simulando un transporte o almacenamiento en refrigeración durante un periodo de tiempo largo. Después de 3 días a 2°C la concentración de CO2 en los envases estuvo entre 1.5—2.6%, mientras que la de O2 fue del 17.8 al 18.9% para las tres superficies de perforación estudiadas. Las concentraciones de los gases permanecieron prácticamente sin cambios durante la primera semana, después cambiaron lentamente hasta que en el día 21 fueron de 7.2—8.8% y 13.6—14.9% para el CO2 y O2, respectivamente. La calidad de la fruta se mantuvo durante los primeros 10 días a 2°C. Después de este periodo aumentó el riesgo de pérdida de calidad, con una marcada pérdida de frutos y desarrollo de sabores indeseables y una pérdida de azúcares y ácidos orgánicos. No hubo prácticamente diferencias entre los envases perforados en cuanto a la calidad de las fresas durante el periodo de 10 días a 2°C, sólo se produjo una ligera pérdida de fruta, no se detectó infección por Botrytis y sólo se detectó una ligera disminución de sacarosa. La comparación con la fruta control durante los 10 primeros días a 2°C no mostró diferencias significativas excepto respecto a los envases que contenían fruta con un contenido de antocianos bajo.

The sodium transporter encoded by the HKT1;2 gene modulates sodium/potassium homeostasis in tomato shoots under salinity

Excessive soil salinity diminishes crop yield and quality. In a previous study in tomato, we identified two closely linked genes encoding HKT1-like transporters, HKT1;1 and HKT1;2, as candidate genes for a major quantitative trait locus (kc7.1) related to shoot Na+ /K+ homeostasis - a major salt tolerance trait - using two populations of recombinant inbred lines (RILs). Here, we determine the effectiveness of these genes in conferring improved salt tolerance by using two near-isogenic lines (NILs) that were homozygous for either the Solanum lycopersicum allele (NIL17) or for the Solanum cheesmaniae allele (NIL14) at both HKT1 loci; transgenic lines derived from these NILs in which each HKT1;1 and HKT1;2 had been silenced by stable transformation were also used. Silencing of ScHKT1;2 and SlHKT1;2 altered the leaf Na+ /K+ ratio and caused hypersensitivity to salinity in plants cultivated under transpiring conditions, whereas silencing SlHKT1;1/ScHKT1;1 had a lesser effect. These results indicate that HKT1;2 has the more significant role in Na+ homeostasis and salinity tolerance in tomato.