Wheat proteomics in the healthgrain project

The project HEALTHGRAIN concerns "Exploiting Bioactivity of European Cereal Grains for Improved Nutrition and Health Benefits". The proteome analysis within HEALTHGRAIN includes establishing the seed proteome reference map of the wheat cultivar Chinese Spring as well as proteome profiling of cultivars selected on the basis of properties relevant for health, nutrition and agronomical performance. The purpose of the proteome mapping is to provide data for integration with genetic analysis and for correlation of protein profiles with specific grain quality properties. In addition individual identified seed proteins will be characterized with regard to spatio-temporal occurrence and post-translational modifications. An overview of the current status of the project focusing on albumins and globulins and tissue localization will be given in addition to an outline of future plans.0info:eu-repo/semantics/publishe

Wheat DOF transcription factors TaSAD and WPBF regulate glutenin gene expression in cooperation with SPA

International audienceGrain storage proteins (GSPs) quantity and composition determine the end-use value of wheat flour. GSPs consists of low-molecular-weight glutenins (LMW-GS), high-molecular-weight glutenins (HMW-GS) and gliadins. GSP gene expression is controlled by a complex network of DNA-protein and protein-protein interactions, which coordinate the tissue-specific protein expression during grain development. The regulatory network has been most extensively studied in barley, particularly the two transcription factors (TFs) of the DNA binding with One Finger (DOF) family, barley Prolamin-box Binding Factor (BPBF) and Scutellum and Aleurone-expressed DOF (SAD). They activate hordein synthesis by binding to the Prolamin box, a motif in the hordein promoter. The BPBF ortholog previously identified in wheat, WPBF, has a transcriptional activity in expression of some GSP genes. Here, the wheat ortholog of SAD, named TaSAD, was identified. The binding of TaSAD to GSP gene promoter sequences in vitro and its transcriptional activity in vivo were investigated. In electrophoretic mobility shift assays, recombinant TaSAD and WPBF proteins bound to cis-motifs like those located on HMW-GS and LMW-GS gene promoters known to bind DOF TFs. We showed by transient expression assays in wheat endosperms that TaSAD and WPBF activate GSP gene expression. Moreover, co-bombardment of Storage Protein Activator (SPA) with WPBF or TaSAD had an additive effect on the expression of GSP genes, possibly through conserved cooperative protein-protein interactions

Wheat DOF transcription factor TaSAD and WPBF regulate glutenin gene expression in cooperation with SPA

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A proteomics survey on wheat susceptibility to Fusarium head blight during grain development.

The mycotoxigenic fungal species Fusarium graminearum is able to attack several important cereal crops, such as wheat and barley. By causing Fusarium Head Blight (FHB) disease, F. graminearum induces yield and quality losses and poses a public health concern due to in planta mycotoxin production. The molecular and physiological plant responses to FHB, and the cellular biochemical pathways used by F. graminearum to complete its infectious process remain still unknown. In this study, a proteomics approach, combining 2D-gel approach and mass spectrometry, has been used to determine the specific protein patterns associated with the development of the fungal infection during grain growth on susceptible wheat. Our results reveal that F. graminearum infection does not deeply alter the grain proteome and does not significantly disturb the first steps of grain ontogeny but impacts molecular changes during the grain filling stage (impact on starch synthesis and storage proteins). The differentially regulated proteins identified were mainly involved in stress and defence mechanisms, primary metabolism, and main cellular processes such as signalling and transport. Our survey suggests that F. graminearum could take advantage of putative susceptibility factors closely related to grain development processes and thus provide new insights into key molecular events controlling the susceptible response to FHB in wheat grains

Storage protein activator controls grain protein accumulation in bread wheat in a nitrogen dependent manner

Abstract The expression of cereal grain storage protein (GSP) genes is controlled by a complex network of transcription factors (TFs). Storage protein activator (SPA) is a major TF acting in this network but its specific function in wheat (Triticum aestivum L.) remains to be determined. Here we generated an RNAi line in which expression of the three SPA homoeologs was reduced. In this line and its null segregant we analyzed GSP accumulation and expression of GSP and regulatory TF genes under two regimes of nitrogen availability. We show that down regulation of SPA decreases grain protein concentration at maturity under low but not high nitrogen supply. Under low nitrogen supply, the decrease in SPA expression also caused a reduction in the total quantity of GSP per grain and in the ratio of GSP to albumin-globulins, without significantly affecting GSP composition. The slight reduction in GSP gene expression measured in the SPA RNAi line under low nitrogen supply did not entirely account for the more significant decrease in GSP accumulation, suggesting that SPA regulates additional levels of GSP synthesis. Our results demonstrate a clear role of SPA in the regulation of grain nitrogen metabolism when nitrogen is a limiting resource

SPA Heterodimerizing Protein, a bZIP transcription factor, represses glutenin synthesis in Triticum aestivum

ISBN: 978-2-9563873-0-5SPA Heterodimerizing Protein, a bZIP transcription factor, represses glutenin synthesis in Triticum aestivum. 2. International Wheat Innovation Worksho

Influence of the allelic variants encoded at the Gli-B1 locus, responsible for a major allergen of wheat, on IgE reactivity for patients suffering from food allergy to wheat

Wheat presents an important genetic diversity that could be useful to look for cultivars with reduced allergencity. omega5-Gliadins have been described as major allergens for wheat allergic patients suffering from wheat-dependent exercise-induced anaphylaxis (WDEIA) and some cases of chronic urticaria (U). Our objective was to study the influence of genetic variability at the Gli-B1 locus encoding for omega5-gliadins on the reactivity of IgE antibodies from these patients. We selected cultivars expressing 13 alleles at Gli-B1 including a wheat/rye translocation and studied the reactivity to gliadins of a rabbit antiserum specific for omega5-gliadins and of IgE from 10 patients. The antiserum and IgE from nine patients with WDEIA and U strongly detected omega5-gliadins expressed by most of the Gli-B1 alleles but showed no or faint responses to the gliadins and secalins extracted from the translocated wheat. The selection of genotypes lacking the Gli-B1 locus may reduce wheat allergenicity

Proteomic and genetic analysis of wheat endosperm albumins and globulins using deletion lines of cultivar Chinese Spring

Albumins and globulins from the endosperm of Triticum aestivum L. cv Chinese Spring (CS) were analysed to establish a proteome reference map for this standard wheat cultivar. Approximately, 1,145 Coomassie-stained spots were detected by two-dimensional gel electrophoresis (2DE), 410 of which were identified using mass spectrometry and data mining. Salt-soluble endosperm proteins from 67 CS deletion lines were also separated by 2DE (four gels per line). Image analysis of the 268 2DE gels as compared to the CS reference proteome allowed the detection of qualitative and quantitative variations in endosperm proteins due to chromosomal deletions. This differential analysis of spots allowed structural or regulatory genes, encoding 211 proteins, to be located on segments of the 21 wheat chromosomes. In addition, variance analysis of quantitative variations in spot volume showed that the expression of 391 proteins is controlled by one or more chromosome bins with 262 significant increases and 196 significant decreases in spot volume. The spot volume of several proteins was increased or decreased by numerous chromosomal regions and homoeologous-like regulation was revealed for some proteins. Quantitative or qualitative variation in a total of 386 proteins was influenced by genes assigned to at least one chromosomal region, while 66 % of all stained proteins were not found to be influenced by chromosome bins. Proteomics of deletion lines can, therefore, be used to simultaneously analyse the composition and genetics of a complex tissue, such as the wheat endosperm.Journal ArticleResearch Support, Non-U.S. Gov'tSCOPUS: ar.jinfo:eu-repo/semantics/publishe

Mapping and proteomic analysis of albumin and globulin proteins in hexaploid wheat kernels (Triticum aestivum L.)

Albumins and globulins of wheat endosperm represent 20% of total kernel protein. They are soluble proteins, mainly enzymes and proteins involved in cell functions. Two-dimensional gel immobiline electrophoresis (2DE) (pH 4-7) x SDS-Page revealed around 2,250 spots. Ninety percent of the spots were common between the very distantly related cultivars 'Opata 85' and 'Synthetic W7984', the two parents of the International Triticeae Mapping Initiative (ITMI) progeny. 'Opata' had 130 specific spots while 'Synthetic' had 96. 2DE and image analysis of the soluble proteins present in 112 recombinant inbred lines of the F9-mapped ITMI progeny enabled 120 unbiased segregating spots to be mapped on 21 wheat (Triticum aestivum L. em. Thell) chromosomes. After trypsic digestion, mapped spots were subjected to MALDI-Tof or tandem mass spectrometry for protein identification by database mining. Among the 'Opata' and 'Synthetic' spots identified, many enzymes have already been mapped in the barley and rice genomes. Multigene families of Heat Shock Proteins, beta-amylases, UDP-glucose pyrophosphorylases, peroxydases and thioredoxins were successfully identified. Although other proteins remain to be identified, some differences were found in the number of segregating proteins involved in response to stress: 11 proteins found in the modern selected cultivar 'Opata 85' as compared to 4 in the new hexaploid 'Synthetic W7984'. In addition, 'Opata' and 'Synthetic' differed in the number of proteins involved in protein folding (2 and 10, respectively). The usefulness of the mapped enzymes for future research on seed composition and characteristics is discussed