Natural Variation in the Content and Degree of Polymerization of Fructans in Wheat: Potential for Selection of Genotypes with Beneficial Health Composition.

Fructans are important biocompounds because of their health-promoting effects as dietary fiber and prebiotics and also because of their harmful effects as fermentable oligosaccharides, disaccharides, monosaccharides, and polyols (FODMAP) particularly in people suffering from irritable bowel syndrome (IBS) and inflammatory bowel disease (IBD), and recently as potential triggers of non-celiac wheat/gluten sensitivity. In this work, we have analyzed the fructan contents as well as its degree of polymerization (DP) in a genetically diverse set of wheat varieties, modern and landraces, from different commonly consumed species (N = 124). A significant variation in fructan contents within and between species was observed, with the following relationship: Triticum aestivum (Landraces) > Triticum aestivum (Modern) ≥ Triticum turgidum (Modern) = T. turgidum (Landraces) ≥ Triticum spelta. In addition, a substantial part of the fructans (>50%) showed a DP ≤ 6. Considering that wheat is a major source of fructans, our results can contribute to a better nutritional management of our diets and be a basis for targeted wheat breeding to alter fructan contents.

Heteroalleles in Common Wheat: Multiple Differences between Allelic Variants of the Gli-B1 Locus.

The Gli-B1-encoded γ-gliadins and non-coding γ-gliadin DNA sequences for 15 different alleles of common wheat have been compared using seven tests: electrophoretic mobility (EM) and molecular weight (MW) of the encoded major γ-gliadin, restriction fragment length polymorphism patterns (RFLPs) (three different markers), Gli-B1-γ-gliadin-pseudogene known SNP markers (Single nucleotide polymorphisms) and sequencing the pseudogene GAG56B. It was discovered that encoded γ-gliadins, with contrasting EM, had similar MWs. However, seven allelic variants (designated from I to VII) differed among them in the other six tests: I (alleles Gli-B1i, k, m, o), II (Gli-B1n, q, s), III (Gli-B1b), IV (Gli-B1e, f, g), V (Gli-B1h), VI (Gli-B1d) and VII (Gli-B1a). Allele Gli-B1c (variant VIII) was identical to the alleles from group IV in four of the tests. Some tests might show a fine difference between alleles belonging to the same variant. Our results attest in favor of the independent origin of at least seven variants at the Gli-B1 locus that might originate from deeply diverged genotypes of the donor(s) of the B genome in hexaploid wheat and therefore might be called "heteroallelic". The donor's particularities at the Gli-B1 locus might be conserved since that time and decisively contribute to the current high genetic diversity of common wheat.

Nitrification inhibitor DMPSA mitigated N2O emission and promoted NO sink in rainfed wheat.

Fertilized cropping systems are important sources of nitrous oxide (N2O) and nitric oxide (NO) to the atmosphere, and biotic and abiotic processes control the production and consumption of these gases in the soil. In fact, the inhibition of nitrification after application of urea or an ammonium-based fertilizer to agricultural soils has resulted in an efficient strategy to mitigate both N2O and NO in aerated agricultural soils. Therefore, the NO and N2O mitigation capacity of a novel nitrification inhibitor (NI), 2-(3,4-dimethyl-1H-pyrazol-1-yl) succinic acid isomeric mixture (DMPSA), has been studied in a winter wheat crop. A high temporal resolution of fluxes of NO and NO2, obtained by using automatic chambers for urea (U) and urea with DMPSA, allowed a better understanding of the temporal net emissions of these gases under field conditions. Seventy-five days after fertilization, the effective reduction of nitrification by DMPSA significantly decreased the production of NO with respect to the treatment without it, giving net consumption of NO in the soil (-61.72 g-N ha-1) for U + DMPSA in comparison to net production (227.44 g-N ha-1) for U. The explanation of NO deposition after NI application, due to biotic and abiotic processes in the soil-plant system, supposes a challenge that needs to be studied in the future. In the case of N2O, the addition of DMPSA significantly mitigated the emissions of this gas by 71%, though the total N2O emissions in both fertilized treatments were significantly greater than those of the control (43.69 g-N ha-1). Regarding the fertilized treatments, no significant effect of DMPSA in comparison to urea alone was observed on grain yield nor bread-making wheat quality. To sum up, we got a significant reduction of N2O and NO with the addition of DMPSA, without a loss in yield and quality parameters in wheat.

In Situ Gluten-Chitosan Interlocked Self-Assembled Supramolecular Architecture Reduces T-Cell-Mediated Immune Response to Gluten in Celiac Disease.

SCOPE: The prevalence of celiac disease has increased since the last half of the 20th century and is now about 1% in most western populations. At present, people who suffer from celiac disease have to follow a gluten-exclusion diet throughout their lives. Compliance to this restrictive diet is demanding and the development of alternative strategies has become urgent. METHODS AND RESULTS: In this context, it is found that the biocompatible aminopolysaccharide chitosan imposes a different gluten reorganization after gluten redox reaction producing in situ mechanically interlocked supramolecular assemblies between gluten and chitosan. These new structures result in the decrease of gluten digestibility, tissue transglutaminase deamidation activity, and interferon-γ production in intestinal T cell lines generated from biopsy specimens of celiac disease patients. CONCLUSION: Overall, the results demonstrate the potential of this research avenue to celiac disease is problematic, as the reorganization of gluten proteins to a novel supramolecular architecture shows a positive impact on known pathogenesis mechanisms of the disease. At present, the only therapy for celiac disease is adherence to a gluten-free diet. Here, it is shown that chitosan-imposed gluten reorganization to an interlocked self-assembled supramolecular architecture reduces gluten digestibility, R5-reactivity, tissue transglutaminase deamidation activity, and its capacity to stimulate a T-cell-mediated immune response in celiac disease.

New insights into wheat toxicity: Breeding did not seem to contribute to a prevalence of potential celiac disease's immunostimulatory epitopes.

Gluten proteins, namely gliadins, are the primary trigger of the abnormal immune response in celiac disease. It has been hypothesised that modern wheat breeding practices may have contributed to the increase in celiac disease prevalence during the latter half of the 20th century. Our results do not support this hypothesis as Triticum aestivum spp. vulgare landraces, which were not subjected to breeding practices, presented higher amounts of potential celiac disease's immunostimulatory epitopes when compared to modern varieties. Furthermore, high variation between wheat varieties concerning the toxic epitopes amount was observed. We carried out quantitative analysis of gliadin types by RP-HPLC to verify its correlation with the amount of toxic epitopes: ω-type gliadins content explain about 40% of the variation of toxic epitopes in tetraploid wheat varieties. This research provides new insights regarding wheat toxicity and into the controversial idea that human practices may have conducted to an increased exposure to toxic epitopes.

Efficient chemo-enzymatic gluten detoxification: reducing toxic epitopes for celiac patients improving functional properties.

Protein engineering of gluten, the exogenous effector in celiac disease, seeking its detoxification by selective chemical modification of toxic epitopes is a very attractive strategy and promising technology when compared to pharmacological treatment or genetic engineering of wheat. Here we present a simple and efficient chemo-enzymatic methodology that decreases celiac disease toxic epitopes of gluten proteins improving its technological value through microbial transglutaminase-mediated transamidation of glutamine with n-butylamine under reducing conditions. First, we found that using low concentrations of amine-nucleophile under non-reducing conditions, the decrease in toxic epitopes is mainly due to transglutaminase-mediated cross-linking. Second, using high amine nucleophile concentrations protein cross-linking is substantially reduced. Third, reducing conditions increase 7-fold the transamidation reaction further decreasing toxic epitopes amount. Fourth, using n-butylamine improves gluten hydrophobicity that strengthens the gluten network. These results open the possibility of tailoring gluten for producing hypoallergenic flours while still taking advantage of the unique viscoelastic properties of gluten.

One hundred years of grain omics: identifying the glutens that feed the world.

Glutens, the storage proteins in wheat grains, are a major source of protein in human nutrition. The protein composition of wheat has therefore been an important focus of cereal research. Proteomic tools have been used to describe the genetic diversity of wheat germplasms from different origins at the level of polymorphisms in alleles encoding glutenin and gliadin, the two main proteins of gluten. More recently, proteomics has been used to understand the impact of specific gluten proteins on wheat quality. Here we review the impact of proteomics on the study of gluten proteins as it has evolved from fractionation and electrophoretic techniques to advanced mass spectrometry. In the postgenome era, proteomics is proving to be essential in the effort to identify and understand the interactions between different gluten proteins. This is helping to fill in gaps in our knowledge of how the technological quality of wheat is determined by the interaction between genotype and environment. We also collate information on the various storage protein alleles identified and their prevalence, which makes it possible to infer the effects of wheat selection on grain protein content. We conclude by reviewing the more recent use of transgenesis aimed at improving the quality of gluten.

Validation of microsatellite markers for cytotype discrimination in the model grass Brachypodium distachyon.

Brachypodium distachyon (L.) P. Beauv. (2n = 2x = 10) is a small annual grass species where the existence of three different cytotypes (10, 20, and 30 chromosomes) has long been regarded as a case of autopolyploid series with x = 5. However, it has been demonstrated that the cytotypes assumed to be polyploids represent two separate Brachypodium species recently named as Brachypodium stacei (2n = 2x = 20) and Brachypodium hybridum (2n = 4x = 30). The aim of this study was to find a PCR-based alternative approach that could replace standard cytotyping methods (i.e., chromosome counting and flow cytometry) to characterize each of the three Brachypodium species. We have analyzed with four microsatellite (SSR) markers 83 B. distachyon-type lines from varied locations in Spain, including the Balearic and Canary Islands. Within this set of lines, 64, 4, and 15 had 10, 20, and 30 chromosomes, respectively. The surveyed markers produced cytotype-specific SSR profiles. So, a single amplification product was generated in the diploid samples, with nonoverlapping allelic ranges between the 2n = 10 and 2n = 20 cytotypes, whereas two bands, one in the size range of each of the diploid cytotypes, were amplified in the 2n = 30 lines. Furthermore, the remarkable size difference obtained with the SSR ALB165 allowed the identification of the Brachypodium species by simple agarose gel electrophoresis.

Down-regulating γ-gliadins in bread wheat leads to non-specific increases in other gluten proteins and has no major effect on dough gluten strength.

BACKGROUND: Gliadins are a major component of gluten proteins but their role in the mixing of dough is not well understood because their contribution to wheat flour functional properties are not as clear as for the glutenin fraction. METHODOLOGY/PRINCIPAL FINDINGS: Transgenic lines of bread wheat with γ-gliadins suppressed by RNAi are reported. The effects on the gluten protein composition and on technological properties of flour were analyzed by RP-HPLC, by sodium dodecyl sulfate sedimentation (SDSS) test and by Mixograph analysis. The silencing of γ-gliadins by RNAi in wheat lines results in an increase in content of all other gluten proteins. Despite the gluten proteins compensation, in silico analysis of amino acid content showed no difference in the γ-gliadins silenced lines. The SDSS test and Mixograph parameters were slightly affected by the suppression of γ-gliadins. CONCLUSIONS/SIGNIFICANCE: Therefore, it is concluded that γ-gliadins do not have an essential functional contribution to the bread-making quality of wheat dough, and their role can be replaced by other gluten proteins.

Genetic variability for waxy genes in Argentinean bread wheat germplasm

Amylose and amylopectin are the two polysaccharides that constitute starch in bread wheat and the enzyme GBSSI (Granule-bound starch synthase I), also known as waxy protein, is responsible for amylose synthesis in storage tissues. Decrease of the amylose content in starch has been associated with the lack of waxy protein(s). In this work, different sets of PCR markers were used to characterize the genetic variability of waxy loci from 103 Argentinean bread wheat cultivars. For the Wx-A1 locus, Wx-A1a and a novel molecular allele designed Wx-A1g were detected. Wx-B1 locus showed three alleles (Wx-B1a, Wx-B1b, Wx-B1e), and Wx-D1 locus showed only the Wx-D1a allele. Novel single-locus allele specific markers for Wx-A1b, Wx-B1b and Wx-D1b null alleles were also described. To our best knowledge this is the first study focused to characterize the genetic variability for waxy genes in bread wheat cultivars from South America.