Selection of GmSWEET39 for oil and protein improvement in soybean.

Soybean [Glycine max (L.) Merr.] was domesticated from wild soybean (G. soja Sieb. and Zucc.) and has been further improved as a dual-use seed crop to provide highly valuable oil and protein for food, feed, and industrial applications. However, the underlying genetic and molecular basis remains less understood. Having combined high-confidence bi-parental linkage mapping with high-resolution association analysis based on 631 whole sequenced genomes, we mapped major soybean protein and oil QTLs on chromosome15 to a sugar transporter gene (GmSWEET39). A two-nucleotide CC deletion truncating C-terminus of GmSWEET39 was strongly associated with high seed oil and low seed protein, suggesting its pleiotropic effect on protein and oil content. GmSWEET39 was predominantly expressed in parenchyma and integument of the seed coat, and likely regulates oil and protein accumulation by affecting sugar delivery from maternal seed coat to the filial embryo. We demonstrated that GmSWEET39 has a dual function for both oil and protein improvement and undergoes two different paths of artificial selection. A CC deletion (CC-) haplotype H1 has been intensively selected during domestication and extensively used in soybean improvement worldwide. H1 is fixed in North American soybean cultivars. The protein-favored (CC+) haplotype H3 still undergoes ongoing selection, reflecting its sustainable role for soybean protein improvement. The comprehensive knowledge on the molecular basis underlying the major QTL and GmSWEET39 haplotypes associated with soybean improvement would be valuable to design new strategies for soybean seed quality improvement using molecular breeding and biotechnological approaches.

Drought stress affects the protein and dietary fiber content of wholemeal wheat flour in wheat/Aegilops addition lines.

Wild relatives of wheat, such as Aegilops spp. are potential sources of genes conferring tolerance to drought stress. As drought stress affects seed composition, the main goal of the present study was to determine the effects of drought stress on the content and composition of the grain storage protein (gliadin (Gli), glutenin (Glu), unextractable polymeric proteins (UPP%) and dietary fiber (arabinoxylan, ß-glucan) components of hexaploid bread wheat (T. aestivum) lines containing added chromosomes from Ae. biuncialis or Ae. geniculata. Both Aegilops parents have higher contents of protein and ß-glucan and higher proportions of water-soluble arabinoxylans (determined as pentosans) than wheat when grown under both well-watered and drought stress conditions. In general, drought stress resulted in increased contents of protein and total pentosans in the addition lines, while the ß-glucan content decreased in many of the addition lines. The differences found between the wheat/Aegilops addition lines and wheat parents under well-watered conditions were also manifested under drought stress conditions: Namely, elevated ß-glucan content was found in addition lines containing chromosomes 5Ug, 7Ug and 7Mb, while chromosomes 1Ub and 1Mg affected the proportion of polymeric proteins (determined as Glu/Gli and UPP%, respectively) under both well-watered and drought stress conditions. Furthermore, the addition of chromosome 6Mg decreased the WE-pentosan content under both conditions. The grain composition of the Aegilops accessions was more stable under drought stress than that of wheat, and wheat lines with the added Aegilops chromosomes 2Mg and 5Mg also had more stable grain protein and pentosan contents. The negative effects of drought stress on both the physical and compositional properties of wheat were also reduced by the addition of these. These results suggest that the stability of the grain composition could be improved under drought stress conditions by the intraspecific hybridization of wheat with its wild relatives.

[Content of Osmolytes and Flavonoids under Salt Stress in Arabidopsis thaliana Plants Defective in Jasmonate Signaling].

The effects of the salt stress (200 mM NaCl) and exogenous jasmonic acid (JA) on levels of osmolytes and flavonoids in leaves of four-week-old Arabidopsis thaliana L. plants of the wild-type (WT) Columbia-0 (Col-0) and the mutant jin1 (jasmonate insensitive 1) with impaired jasmonate signaling were studied. The increase in proline content caused by the salt stress was higher in the Col-0 plants than in the mutant jin1. This difference was especially marked if the plants had been pretreated with exogenous 0.1 µM JA. The sugar content increased in response to the salt stress in the JA-treated WT plants but decreased in the jin1 mutant. Leaf treatment with JA of the WT plants but not mutant defective in jasmonate signaling also enhanced the levels of anthocyanins and flavonoids absorbed in UV-B range. The presence of JA increased salinity resistance of the Col-0 plants, since the accumulation of lipid peroxidation products and growth inhibition caused by NaCl were less pronounced. Under salt stress, JA almost did not render a positive effect on the jin1 plants. It is concluded that the protein JIN1/MYC2 is involved in control of protective systems under salt stress.

Chalky part differs in chemical composition from translucent part of japonica rice grains as revealed by a notched-belly mutant with white-belly.

BACKGROUND: Chalkiness has a deleterious influence on rice appearance and milling quality. We identified a notched-belly mutant with a high percentage of white-belly, and thereby developed a novel comparison system that can minimize the influence of genetic background and growing conditions. Using this mutant, we examined the differences in chemical composition between chalky and translucent endosperm, with the aim of exploring relations between occurrence of chalkiness and accumulation of starch, protein and minerals. RESULTS: Comparisons showed a significant effect of chalkiness on chemical components in the endosperm. In general, occurrence of chalkiness resulted in higher total starch concentration and lower concentrations of the majority of the amino acids measured. Chalkiness also had a positive effect on the concentrations of As, Ba, Cd, Cr, Mn, Na, Sr and V, but was negatively correlated with those of B, Ca, Cu, Fe and Ni. By contrast, no significant chalkiness effect on P, phytic acid-P, K, Mg or Zn was observed. In addition, substantial influence of the embryo on endosperm composition was detected, with the embryo showing a negative effect on total protein, amino acids such as Arg, His, Leu, Lys, Phe and Tyr, and all the 17 minerals measured, excluding Ca, Cu, P and Sr. CONCLUSION: An inverse relation between starch and protein as well as amino acids was found with respect to chalkiness occurrence. Phytic acid and its colocalized elements K and Mg were not affected by chalkiness. The embryo exerted a marked influence on chemical components of the endosperm, in particular minerals, suggesting the necessity of examining the role of the embryo in chalkiness formation. © 2016 The Authors. Journal of the Science of Food and Agriculture published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

Nutritional composition, bioactive compounds and volatile profile of cocoa beans from different regions of Cameroon.

Analysis of the complex composition of cocoa beans provides fundamental information for evaluating the quality and nutritional aspects of cocoa-based food products, nutraceuticals and supplements. Cameroon, the world's fourth largest producer of cocoa, has been defined as "Africa in miniature" because of the variety it habitats. In order to evaluate the nutritional characteristics of cocoa beans from five different regions of Cameroon, we studied their polyphenolic content, volatile compounds and fatty acids composition. The High Performance Thin Layer Chromatography (HPTLC) analysis showed that the Mbalmayo sample had the highest content of theobromine (11.6 mg/g) and caffeic acid (2.1 mg/g), while the Sanchou sample had the highest level of (-)-epicatechin (142.9 mg/g). Concerning fatty acids, the lowest level of stearic acid was found in the Mbalmayo sample while the Bertoua sample showed the highest content of oleic acid. Thus, we confirmed that geographical origin influences the quality and nutritional characteristics of cocoa from these regions of Cameroon.

Tailoring Grain Storage Reserves for a Healthier Rice Diet and its Comparative Status with Other Cereals.

A global rise of diet-related noncommunicable diseases calls for a focus on diet-based nutritional intervention across the entire socioeconomic consumer spectrum. We review recent reports in the area of healthier rice aimed at developing rice grains with improved dietary fiber compositions (increased amounts of nonstarch polysaccharides and resistant starch), and less digestible starch (higher amylose and phospholipid complex in the endosperm) resulting in reduced glycemic impact upon grain consumption. We furthermore elaborate on the interconnections of elevated amounts of protein and a balanced composition of essential amino acids. The importance of a nutritious aleurone layer and its role in lipid storage and micronutrient composition is discussed briefly in the context of brown rice benefits. We identify gene targets for precision breeding that will facilitate the production of rice grains and rice-based products to mitigate the impact of nutrition-related preventable diseases.

Tuber proteome comparison of five potato varieties by principal component analysis.

BACKGROUND: Data analysis of omics data should be performed by multivariate analysis such as principal component analysis (PCA). The way data are clustered in PCA is of major importance to develop some classification systems based on multivariate analysis, such as soft independent modeling of class analogy (SIMCA). In a previous study a one-class classifier based on SIMCA was built using microarray data from a set of potatoes. The PCA grouped the transcriptomic data according to varieties. The present work aimed to use PCA to verify the clustering of the proteomic profiles for the same potato varieties. RESULTS: Proteomic profiles of five potato varieties (Biogold, Fontane, Innovator, Lady Rosetta and Maris Piper) were evaluated by two-dimensional gel electrophoresis (2-DE) performed on two immobilized pH gradient (IPG) strip lengths, 13 and 24 cm, both under pH range 4-7. For each strip length, two gels were prepared from each variety; in total there were ten gels per analysis. For 13 cm strips, 199-320 spots were detected per gel, and for 24 cm strips, 365-684 spots. CONCLUSION: All four PCAs performed with these datasets presented clear grouping of samples according to the varieties. The data presented here showed that PCA was applicable for proteomic analysis of potato and was able to separate the samples by varieties. © 2016 Society of Chemical Industry.

Effect of germination time on proximate analysis, bioactive compounds and antioxidant activity of lentil (Lens culinaris Medik.) sprouts.

BACKGROUND: The lentil plant, Lens culinaris L., is a member of the Leguminoceae family and constitutes one of the most important traditional dietary components. The purpose of the current study was to investigate the effects of sprouting for 3, 4, 5 and 6 days on proximate, bioactive compounds and antioxidative characteristics of lentil (Lens culinaris) sprouts. MATERIAL AND METHODS: Lentil seeds were soaked in distilled water (1:10, w/v) for 12 h at room temperature (~25°C), then kept between thick layers of cotton cloth and allowed to germinate in the dark for 3, 4, 5 and 6 days. The nutritional composition, protein solubility, free amino acids, antinutritional factors, bioactive compounds and antioxidant activity of raw and germinated samples were determined using standard official procedures. RESULTS: Sprouting process caused significant (P ≤ 0.05) increases in moisture, protein, ash, crude fiber, protein solubility, free amino acids, total, reducing and nonreducing sugars. However, oil content, antinutritional factors (tannins and phytic acid) significantly (P ≤ 0.05) decreased. Results indicated that total essential amino acids of lentil seeds protein formed 38.10% of the total amino acid content. Sulfur-containing amino acids were the first limiting amino acid, while threonine was the second limiting amino acid in raw and germinated lentil seeds. Sprouting process has a positive effect on the essential amino acid contents and protein efficiency ratio (PER) of lentil sprouts. Phenolics content increased from 1341.13 mg/100 g DW in raw lentil seeds to 1411.50, 1463.00, 1630.20 and 1510.10 in those samples germinated for 3, 4, 5 and 6 days, respectively. Sprouted seeds had higher DPPH radical scavenging and reducing power activities. CONCLUSIONS: Based on these results, sprouting process is recommended to increase nutritive value, and antioxidant activity of lentil seeds.

Isolation of salt stress-related genes from Aspergillus glaucus CCHA by random overexpression in Escherichia coli.

The halotolerant fungus Aspergillus glaucus CCHA was isolated from the surface of wild vegetation around a saltern with the salinity range being 0-31%. Here, a full-length cDNA library of A. glaucus under salt stress was constructed to identify genes related to salt tolerance, and one hundred clones were randomly selected for sequencing and bioinformatics analysis. Among these, 82 putative sequences were functionally annotated as being involved in signal transduction, osmolyte synthesis and transport, or regulation of transcription. Subsequently, the cDNA library was transformed into E. coli cells to screen for putative salt stress-related clones. Five putative positive clones were obtained from E. coli cells grown on LB agar containing 1 M NaCl, on which they showed rapid growth compared to the empty vector control line. Analysis of transgenic Arabidopsis thaliana lines overexpressing CCHA-2142 demonstrated that the gene conferred increased salt tolerance to plants as well by protecting the cellular membranes, suppressing the inhibition of chlorophyll biosynthesis. These results highlight the utility of this A. glaucus cDNA library as a tool for isolating and characterizing genes related to salt tolerance. Furthermore, the identified genes can be used for the study of the underlying biology of halotolerance.

Effect of NaCl stress on health-promoting compounds and antioxidant activity in the sprouts of three broccoli cultivars.

Health-promoting compounds, antioxidant and myrosinase activity in the sprouts of three broccoli cultivars under 40 mM, 80 mM and 160 mM NaCl were investigated. LangYan (LY) sprouts had the richest health-promoting compounds among the evaluated cultivars. Treatment of 40 mM and 80 mM NaCl significantly decreased the content of ascorbic acid and total phenolic as well as antioxidant activity, but did not affect glucoraphanin, sulforaphane and myrosinase activity compared to the control. However, 160 mM NaCl treatment significantly enhanced the level of total phenolic, glucoraphanin, sulforaphane, antioxidant and myrosinase activity, while significantly decreased ascorbic acid content. The results suggest that 160 mM NaCl treatment would enhance antioxidant activity and sulforaphane yield in broccoli sprouts. The health-promoting value of broccoli sprouts depends on plant genotype and could be affected by NaCl stress.

Metabolic and bioprocess engineering for production of selenized yeast with increased content of seleno-methylselenocysteine.

Specific Se-metabolites have been recognized to be the main elements responsible for beneficial effects of Se-enriched diet, and Se-methylselenocysteine (SeMCys) is thought to be among the most effective ones. Here we show that an engineered Saccharomyces cerevisiae strain, expressing a codon optimized heterologous selenocysteine methyltransferase and endowed with high intracellular levels of S-adenosyl-methionine, was able to accumulate SeMCys at levels higher than commercial selenized yeasts. A fine tuned carbon- and sulfate-limited fed-batch bioprocess was crucial to achieve good yields of biomass and SeMCys. Through the coupling of metabolic and bioprocess engineering we achieved a ∼24-fold increase in SeMCys, compared to certified reference material of selenized yeast. In addition, we investigated the interplay between sulfur and selenium metabolism and the possibility that redox imbalance occurred along with intracellular accumulation of Se. Collectively, our data show how the combination of metabolic and bioprocess engineering can be used for the production of selenized yeast enriched with beneficial Se-metabolites.

Cloning, expression, characterization, and computational approach for cross-reactivity prediction of manganese superoxide dismutase allergen from pistachio nut.

BACKGROUND: Tree nut allergy is one of the common potentially life-threatening food allergies in children and adults. Recombinant food allergens offer new perspectives to solve problems of clinical and molecular allergology in diagnosis, research, and therapy of food allergies. So far, superoxide dismutase (s) has been identified as a panallergen and studied in different allergenic sources. Manganese Superoxide Dismutase (MnSOD) has also been reported in pistachio that may cause allergic reactions in atopic subjects. The aim of this study was to describe the cloning, expression, and purification of MnSOD from pistachio nut. METHODS: The pistachio MnSOD was cloned and expressed in E. coli BL21 (DE3) using a vector pET-32b (+). A recombinant protein was purified by metal precipitation. The protein immunoreactivity was evaluated using patients' IgE binding by means of ELISA and immunoblotting assays. RESULTS: The MnSOD gene from pistachio was successfully cloned and expressed in E. coli. The purified pistachio MnSOD was recognized by IgE in 10 (40%) out of the 25 sera tested. Our results also showed that this protein might trigger some cross-reactions toward IgE antibodies and thus could be considered as a panallergen. CONCLUSIONS: For the first time recombinant manganese superoxide dismutase from nut source was expressed as a possible allergen. This pistachio allergen could be a possible basis for cross-reactivity with MnSOD from other sources.

Bioconversion of low quality lignocellulosic agricultural waste into edible protein by Pleurotus sajor-caju (Fr.) Singer.

Pleurotus sajor-caju (Fr.) Singer was cultivated on selected agro wastes viz. cotton stalks, groundnut haulms, soybean straw, pigeon pea stalks and leaves and wheat straw, alone or in combinations. Cotton stalks, pigeon pea stalks and wheat straw alone or in combination were found to be more suitable than groundnut haulms and soybean straw for the cultivation. Organic supplements such as groundnut oilseed cake, gram powder and rice bran not only affected growth parameters but also increased yields. Thus bioconversion of lignocellulosic biomass by P. sajor-caju offers a promising way to convert low quality biomass into an improved human food.

Subcellular localization of the 2S globulin narbonin in seeds of Vicia narbonensis.

Narbonin is a 2S protein from the globulin fraction of narbon bean (Vicia narbonensis L.) cotyledons. Its amino acid composition and the pattern of its regulated accumulation in developing seeds led to the suggestion that narbonin could be a storage protein. Therefore, it was expected to be present in protein bodies of the storage tissue cells. Comparison of the cDNA-derived amino acid sequence with a directly determined partial N-terminal sequence revealed that the primary translation product of narbonin mRNA lacks a transient N-terminal signal peptide (V.H. Nong et al., 1995, Plant Mol Biol 28: 61 - 72). Narbonin polypeptides that had been synthesized in a cell-free translation system supplemented with dog pancreas microsomes were not protected against degradation by posttranslationally added proteases (protease protection assay). In accordance with the lack of a signal peptide this indicates that the polypeptide was not cotranslationally sequestered into the microsomes. The protein-body fraction that had been isolated from mature narbon bean cotyledons by a non-aqueous gradient centrifugation procedure was free of narbonin; this was found in the soluble cell fraction. In electron micrographs, narbonin could be localized in the cytoplasm using the immuno gold-labelling technique. Previously, it had already been shown that narbonin is too slowly degraded during narbon bean germination to act as a storage protein. From all these results it has to be concluded that narbonin is a cytoplasmic protein which does not belong to the storage proteins in the restricted sense. Other possible functions are discussed.

Developmental and environmental regulation of pea legumin genes in transgenic tobacco.

Two distinct legumin genes (LegA1 and LegA2) which encode a major class of seed storage protein in pea were isolated from a genomic library. The cloned fragments were introduced into tobacco via Agrobacterium-mediated transformation and the regenerated plants were used to study the expression characteristics of the genes in a heterologous host. It was found that both LegA1 and LegA2 were functional members of the pea legumin gene family and that their expression was similar in both pea and transgenic tobacco. Legumin was detected only in the seed of tobacco where the primary translation products were processed in a manner analogous to that which occurs in pea. Legumin gene expression was also shown to be temporally regulated during seed development. Legumin polypeptides and mRNA began to accumulate 16 days after flowering (DAF), in contrast to the endogenous tobacco storage proteins which were apparent at 13 DAF. It was also demonstrated that the legumin genes in tobacco were environmentally regulated to the nutritional status of the plant. As has been previously shown in pea, legumin accumulation in transgenic tobacco seed was progressively reduced when the plants were grown under conditions of increasing severity of sulphur-nutrient stress. The reduced accumulation of protein was correlated with lower levels of legumin mRNA in the developing seed. Despite encoding nearly identical subunits, nucleotide sequence data for LegA1 and LegA2 showed that the similarity of their respective 5'-flanking regions was restricted to several short elements mostly within 240 bp from the start of transcription. However, a deletion series using the LegA1 gene demonstrated that 237 bp of 5'-flanking sequence was insufficient to permit the expression of the legumin gene in tobacco. The data indicated that an as yet unidentified sequence element(s) located between positions -668 and -237 was essential in re-establishing the high level of regulated gene expression observed with the full-length LegA1 gene.

Restoration of light induced photosystem II inhibition without de novo protein synthesis.

Illumination of isolated spinach thylakoid membranes under anaerobic conditions gave rise to severe inhibition of photosystem II electron transport but did not result in D1-protein degradation. When these photoinhibited thylakoids were incubated in total darkness the photosystem II activity could be fully restored in vitro in a process that required 1-2 h for completion.

The effect of different alleles at the r locus on the synthesis of seed storage proteins in Pisum sativum.

Rocket immunoelectrophoresis was used to measure the accumulation of storage proteins in developing cotyledons of two Pisum sativum (pea) genotypes, that were close to isogenic except for the nature of the allele at the r locus. There was a marked decrease in legumin accumulation in the rr (wrinkled-seeded) genotype compared to the RR (round-seeded) genotype. The accumulation of vicilin did not differ greatly between the two genotypes. Pulse-labelling studies indicated that the differences in rates of accumulation of legumin between the rr and RR genotypes were a consequence of differences in rates of protein synthesis. Measurements of relative amounts of specific mRNAs, using cDNA clones as probes, showed lower amounts of legumin mRNA in developing cotyledons of the rr, compared to the RR, genotype. Both vicilin mRNAs and convicilin mRNA, the latter of which shows a similar temporal pattern of expression to those of the major legumin species, are relatively unaffected by the nature of the allele at the r locus. Nuclear run-on transcription experiments indicated no differences in the rate of synthesis of legumin transcripts in the rr and RR near-isolines. The consequences of homozygosity for the r allele on storage protein mRNA levels in vitro may be mimicked by manipulating the sucrose concentration of the culture medium.

Isolation and expression of a pea vicilin cDNA in the yeast Saccharomyces cerevisiae.

A cDNA clone containing the complete coding sequence for vicilin from pea (Pisum sativum L.) was isolated. It specifies a 50,000-Mr protein that in pea is neither post-translationally processed nor glycosylated. The cDNA clone was expressed in yeast from a 2 micron plasmid by using the yeast phosphoglycerate kinase promoter and initiator codon. The resultant fusion protein, which contains the first 16 amino acid residues of phosphoglycerate kinase in addition to the vicilin sequence, was purified and subsequently characterized. It has slightly slower mobility on SDS/polyacrylamide-gel electrophoresis than standard pea vicilin and forms a mixture of multimers, some of which resemble the native protein.

Regulation of storage-protein synthesis in pea (Pisum sativum L.) cotyledons under conditions of sulphur deficiency.

The effects of sulphur deficiency on the expression of storage-protein genes in developing pea (Pisum sativum) cotyledons were studied. Legumin-gene transcription was decreased by S-deficiency, but not to the same extent as the decrease in the level of legumin mRNA. Vicilin-gene transcription was not significantly affected. Control of gene expression may thus occur during transcription and/or post-transcriptional events.