Seed Storage Proteins Research Articles

Integrated ATAC-Seq and RNA-Seq Data Analysis to Reveal OsbZIP14 Function in Rice in Response to Heat Stress.

Transcription factors (TFs) play critical roles in mediating the plant response to various abiotic stresses, particularly heat stress. Plants respond to elevated temperatures by modulating the expression of genes involved in diverse metabolic pathways, a regulatory process primarily governed by multiple TFs in a networked configuration. Many TFs, such as WRKY, MYB, NAC, bZIP, zinc finger protein, AP2/ERF, DREB, ERF, bHLH, and brassinosteroids, are associated with heat shock factor (Hsf) families, and are involved in heat stress tolerance. These TFs hold the potential to control multiple genes, which makes them ideal targets for enhancing the heat stress tolerance of crop plants. Despite their immense importance, only a small number of heat-stress-responsive TFs have been identified in rice. The molecular mechanisms underpinning the role of TFs in rice adaptation to heat stress still need to be researched. This study identified three TF genes, including OsbZIP14, OsMYB2, and OsHSF7, by integrating transcriptomic and epigenetic sequencing data analysis of rice in response to heat stress. Through comprehensive bioinformatics analysis, we demonstrated that OsbZIP14, one of the key heat-responsive TF genes, contained a basic-leucine zipper domain and primarily functioned as a nuclear TF with transcriptional activation capability. By knocking out the OsbZIP14 gene in the rice cultivar Zhonghua 11, we observed that the knockout mutant OsbZIP14 exhibited dwarfism with reduced tiller during the grain-filling stage. Under high-temperature treatment, it was also demonstrated that in the OsbZIP14 mutant, the expression of the OsbZIP58 gene, a key regulator of rice seed storage protein (SSP) accumulation, was upregulated. Furthermore, bimolecular fluorescence complementation (BiFC) experiments uncovered a direct interaction between OsbZIP14 and OsbZIP58. Our results suggested that OsbZIP14 acts as a key TF gene through the concerted action of OsbZIP58 and OsbZIP14 during rice filling under heat stress. These findings provide good candidate genes for genetic improvement of rice but also offer valuable scientific insights into the mechanism of heat tolerance stress in rice.

Large and stable genome edits at the sorghum alpha kafirin locus result in changes in chromatin accessibility and globally increased expression of genes encoding lysine enrichment.

Sorghum is a resilient and widely cultivated grain crop used for feed and food. However, it's grain is deficient in lysine, an essential amino acid. This is due to the primary seed storage proteins, the alpha-kafirins, lacking lysine. It has been observed that reductions in alpha-kafirin protein results in rebalancing of the seed proteome and a corresponding increase in non-kafirin proteins which leads to an increased lysine content. However, the mechanisms underlying proteome rebalancing are unclear. This study characterizes a previously developed gene edited sorghum line, with deletions at the alpha kafirin locus. A single consensus guide RNA leads to tandem deletion of multiple members of the gene family in addition to the small target site mutations in remaining genes. RNA-seq and ATAC-seq were utilized to identify changes in gene expression and chromatin accessibility in developing kernels in the absence of most alpha-kafirin expression. Several differentially accessible chromatin regions and differentially expressed genes were identified. Additionally, several genes upregulated in the edited sorghum line were common with their syntenic orthologues differentially expressed in maize prolamin mutants. ATAC-seq showed enrichment of the binding motif for ZmOPAQUE 11, perhaps indicating the transcription factor's involvement in the kernel response to reduced prolamins. Overall, this study provides a resource of genes and chromosomal regions which may be involved in sorghum's response to reduced seed storage proteins and the process of proteome rebalancing.

Retrograde transport in plants: Circular economy in the endomembrane system

The study of endomembrane trafficking is crucial for understanding how cells and whole organisms function. Moreover, there is a special interest in investigating endomembrane trafficking in plants, given its role in transport and accumulation of seed storage proteins and in secretion of cell wall material, arguably the two most essential commodities obtained from crops. The mechanisms of anterograde transport in the biosynthetic and endocytic pathways of plants have been thoroughly discussed in recent reviews, but, comparatively, retrograde trafficking pathways have received less attention. Retrograde trafficking is essential to recover membranes, retrieve proteins that have escaped from their intended localization, maintain homeostasis in maturing compartments, and recycle trafficking machinery for its reuse in anterograde transport reactions. Here, we review the current understanding on retrograde trafficking pathways in the endomembrane system of plants, discussing their integration with anterograde transport routes, describing conserved and plant-specific retrieval mechanisms at play, highlighting contentious issues and identifying open questions for future research.

Estimation of genetic diversity using seed storage protein (SSP) profiling in wild and cultivated species of Cicer L

The narrow genetic diversity of chickpea is a serious impediment to modern cultivar creation. Seed storage proteins (SSPs) are stable and have minimal or no degradation when subjected to isolation and SDS-PAGE. We have characterized SSPs of 436 chickpea genotypes, belonging to nine annual Cicer species, originated from 47 countries by SDS-PAGE and determined the extent of genetic diversity in chickpea through clustering. Based on scoring, a total of 44 bands (10 to 170kDa) were identified, which were all polymorphic. The least appeared protein bands were 11, 160 and 170kDa where band of 11 and 160kDa was present exclusively in wild type. Five bands were present in < 10% of genotypes. Bands appeared in 200-300 genotypes were suggested less polymorphic, on contrary bands present in 10-150 genotypes were suggested more polymorphic. Polymorphism of protein bands in context to their potential functions reported in literature were explored and suggested that the glubulins were most and glutelins were least abundant, whereas albumins with their known role in stress tolerance can be used as marker in chickpea breeding. Cluster analysis produced 14 clusters, interestingly three clusters contained only Pakistani genotypes and thus Pakistani genotypes appeared as a separate entity from the rest of the genotypes. Our results indicate that SDS-PAGE of SSPs is a powerful technique in determining the genetic diversity plus it is easily adaptable, due to its cost effectiveness in comparison to other genomics tools.

Morphology, Formation Kinetics and Core Composition of Pea and Soy 7S and 11S Globulin Amyloid Fibrils.

Legume seed storage proteins can be induced to form amyloid fibrils upon heating at low pH, which could improve their functionality for use in food and materials. However, the amyloidogenic regions of legume proteins are largely unknown. Here, we used LC-MS/MS to determine the amyloid core regions of fibrils formed by enriched pea and soy 7S and 11S globulins at pH 2, 80 °C, and characterized their hydrolysis, assembly kinetics, and morphology. A lag phase was absent from the fibrillation kinetics of pea and soy 7S globulins, while 11S globulins and crude extracts displayed a similar lag time. Pea and soy protein fibrils differed in morphology, with most pea fibrils being straight and soy fibrils being worm-like. Pea and soy globulins were abundant in amyloid-forming peptides, with over 100 unique fibril-core peptides from pea 7S and around 50 unique fibril-core peptides identified from pea 11S, soy 7S, and soy 11S globulins. Amyloidogenic regions derive predominantly from the homologous core region of 7S globulins and the basic subunit of 11S globulins. Overall, pea and soy 7S and 11S globulins are rich in amyloidogenic regions. This study will help understand their fibrillation mechanism and engineer protein fibrils with specific structures and functionality.

Influence on Accumulation Levels and Subcellular Localization of Prolamins by Fusion with the Functional Peptide in Transgenic Rice Seeds.

To exploit the rice seed-based oral vaccine against Sjögren's syndrome, altered peptide ligand of N-terminal 1 (N1-APL7) from its M3 muscarinic acetylcholine receptor (M3R) autoantigen was expressed as fusion protein with the representative four types of rice prolamins (16kDa, 14kDa, 13kDa, and 10kDa prolamins) under the control of the individual native prolamin promoter. The 10kD:N1-APL7 and 14kD:N1-APL7 accumulated at high levels (287 and 58µg/grain), respectively, whereas production levels of the remaining ones were remarkably low. Co-expression of these fusion proteins did not enhance the accumulation level of N1-APL7 in an additive manner. Downregulation of endogenous seed storage proteins by RNAi-mediated suppression also did not lead to substantial elevation of the co-expressed prolamin:N1-APL7 products. When transgenic rice seeds were subjected to in vitro proteolysis with pepsin, the 10kD:N1-APL7 was digested more quickly than the endogenous 10kDa prolamin and the 14kD:N1-APL7 deposited in PB-Is. This difference could be explained by the finding that the 10kD:N1-APL7 was unexpectedly localized in the PB-IIs containing glutelins. These results indicated that not only accumulation level but also subcellular localization of inherent prolamins were highly influenced by the liked N1-APL7 peptide.

Wheat quality: A review on chemical composition, nutritional attributes, grain anatomy, types, classification, and function of seed storage proteins in bread making quality.

Wheat (Triticum aestivum L.) belonging to one of the most diverse and substantial families, Poaceae, is the principal cereal crop for the majority of the world's population. This cereal is polyploidy in nature and domestically grown worldwide. Wheat is the source of approximately half of the food calories consumed worldwide and is rich in proteins (gluten), minerals (Cu, Mg, Zn, P, and Fe), vitamins (B-group and E), riboflavin, niacin, thiamine, and dietary fiber. Wheat seed-storage proteins represent an important source of food and energy and play a major role in the determination of bread-making quality. The two groups of wheat grain proteins, i.e., gliadins and glutenins, have been widely studied using SDS-PAGE and other techniques. Sustainable production with little input of chemicals along with high nutritional quality for its precise ultimate uses in the human diet are major focus areas for wheat improvement. An expansion in the hereditary base of wheat varieties must be considered in the wheat breeding program. It may be accomplished in several ways, such as the use of plant genetic resources, comprising wild relatives and landraces, germplasm-assisted breeding through advanced genomic tools, and the application of modern methods, such as genome editing. In this review, we critically focus on phytochemical composition, reproduction growth, types, quality, seed storage protein, and recent challenges in wheat breeding and discuss possible ways forward to combat those issues.

Proteomic Variability and Nutrient-Related Proteins across Pigmented and Non-Pigmented Rice Grains

Rice protein is considered an important dietary protein source. Information regarding rice nutrient-related protein expression is still scarce, hence further study investigating this aspect is highly needed. Herein, we applied sequential window acquisition of all theoretical mass spectra (SWATH-MS) for a comparative proteomic analysis across six different Malaysian rice varieties. These consisted of black rice (BR: PH9 and BALI), red rice (RR: MRQ100 and MRM16), and white rice (WR: MRQ76 and MR297). This study aimed to unravel rice nutrient-related proteins and if their expressions were significantly different across varieties. A total of 4022 quantified proteins were found to be significantly expressed across all varieties with a false discovery rate (FDR) &lt; 1% and p &lt; 0.05. While among 1792 differentially expressed proteins (DEPs) that were identified, 74 DEPs had functions related to nutrient biosynthesis. There were significantly higher expressions of key enzymes for the carotenoid and amylopectin biosynthesis pathways and seed storage proteins, i.e., prolamins and glutelins in RR. Glycoproteins such as cupin and germin-like protein, as well as enzymes that are involved in the biosynthesis of thiamine and anthocyanin were abundantly found in BR. WR was particularly enriched with biosynthesis enzymes for essential amino acids (methionine and arginine), vitamin B, and unsaturated fatty acid. This study provides us insights into the differential expressions of storage and functional proteins with nutrient-related properties in shaping rice grain pigmentations and plant immunity, as well as in contributing diverse health benefits as daily functional food for human consumption.

Cross-Serological Reaction of Glandless Cottonseed Proteins to Peanut and Tree Nut Allergic IgE

Food allergy is a potentially life-threatening health concern caused by immunoglobulin E (IgE) antibodies that mistakenly recognize normally harmless food proteins as threats. Peanuts and tree nuts contain several seed storage proteins that commonly act as allergens. Glandless cottonseed, lacking the toxic compound gossypol, is a new food source. However, the seed storage proteins in cottonseed may act as allergens. To assess this risk, glandless cottonseed protein extracts were evaluated for IgE binding by peanut and tree nut allergic volunteers. ELISA demonstrated that 25% of 32 samples had significant binding to cottonseed extracts. Immunoblot analysis with pooled sera indicated that IgE recognized a pair of bands migrating at approximately 50 kDa. Excision of these bands and subsequent mass-spectrometric analysis demonstrated peptide matches to cotton C72 and GC72 vicilin and legumin A and B proteins. Further, in silico analysis indicated similarity of the cotton vicilin and legumin proteins to peanut vicilin (Ara h 1) and cashew nut legumin (Ana o 2) IgE-binding epitopes among others. The observations suggest both the cotton vicilin and legumin proteins were recognized by the nut allergic IgE, and they should be considered for future allergen risk assessments evaluating glandless cottonseed protein products.

Development and Optimization of Label-Free Quantitative Proteomics under Different Crossing Periods of Bottle Gourd.

Bottle gourd, a common vegetable in the human diet, has been valued for its medicinal and energetic properties. In this experiment, the time-resolved analysis of the changes in the proteins' electrophoretic patterning of the seed development at different crossing periods was studied in bottle gourd using label-free quantitative proteomics. Hybrid HBGH-35 had the highest observed protein levels at the 4th week of the crossing period (F4) compared to the parental lines, viz. G-2 (M) and Pusa Naveen (F). The crossing period is significantly correlated with grain filling and reserve accumulation. The observed protein expression profile after storage was related to seed maturation and grain filling in bottle gourds. A total of 2517 proteins were identified in differentially treated bottle gourd fruits, and 372 proteins were differentially expressed between different crossing periods. Proteins related to carbohydrate and energy metabolism, anthocyanin biosynthesis, cell stress response, and fruit firmness were characterized and quantified. Some proteins were involved in the development, while others were engaged in desiccation and the early grain-filling stage. F4 was distinguished by an increase in the accumulation of low molecular weight proteins and enzymes such as amylase, a serine protease, and trypsin inhibitors. The seed vigor also followed similar patterns of differential expression of seed storage proteins. Our findings defined a new window during seed production, which showed that at F4, maximum photosynthetic assimilates accumulated, resulting in an enhanced source-sink relationship and improved seed production. Our study attempts to observe the protein expression profiling pattern under different crossing periods using label-free quantitative proteomics in bottle gourd. It will facilitate future detailed investigation of the protein associated with quality traits and the agronomic importance of bottle gourd through selective breeding programs.

Introgression of Seedling Plant Resistance to Leaf Rust from Agropyron cristatum into Wheat by Induced Homoeologous Recombination

Agropyron cristatum (P genome) is a Triticeae species from the wheat tertiary gene pool which has economic importance as forage and also displays traits beneficial to wheat. Resistance to leaf rust was previously mapped to the short arm of chromosome 1P (1PS) in A. cristatum by the development of a compensating Robertsonian translocation involving chromosome arm 1PS and the long arm of wheat chromosome 1B (1BL). In this study, chromosome arm 1PS was engineered using the ph1b mutation to induce 1BS/1PS homoeologous recombination and to obtain new translocations with shortened fragments of chromosome arm 1PS. Two translocations with different alien fragment sizes were identified by genomic in situ hybridization, wheat 1BS- and 1PS-specific molecular markers and gene-specific markers for glutenin, Glu-B3 and gliadin Gli-B1 seed storage protein. One translocation (called type 1) replaces a proximal segment of 1PS chromatin, and the other (called type 2) replaces a distal 1PS segment and introduces the Glu-B3 and Gli-B1 wheat storage protein loci. Six specific EST-STS markers for chromosome arm 1PS amplified PCR products in the recombinant type 2 translocation line. Resistance analysis showed that the type 2 translocation was highly resistant to a virulent race of leaf rust pathogen. The new wheat–A. cristatum translocations obtained yield material with seedling plant resistance to leaf rust and seed storage protein loci.

Seed Storage Protein, Functional Diversity and Association with Allergy

Plants are essential for humans as they serve as a source of food, fuel, medicine, oils, and more. The major elements that are utilized for our needs exist in storage organs, such as seeds. These seeds are rich in proteins, show a broad spectrum of physiological roles, and are classified based on their sequence, structure, and conserved motifs. With the improvements to our knowledge of the basic sequence and our structural understanding, we have acquired better insights into seed proteins and their role. However, we still lack a systematic analysis towards understanding the functional diversity associated within each family and their associations with allergy. This review puts together the information about seed proteins, their classification, and diverse functional roles along with their associations with allergy.

Transcriptome and Proteome Analysis Revealed the Influence of High-Molecular-Weight Glutenin Subunits (HMW-GSs) Deficiency on Expression of Storage Substances and the Potential Regulatory Mechanism of HMW-GSs.

The processing quality of wheat is affected by seed storage substances, such as protein and starch. High-molecular-weight glutenin subunits (HMW-GSs) are the major components of wheat seed storage proteins (SSPs); they are also key determinators of wheat end-use quality. However, the effects of HMW-GSs absence on the expression of other storage substances and the regulation mechanism of HMW-GSs are still limited. Previously, a wheat transgenic line LH-11 with complete deletions of HMW-GSs was obtained through introducing an exogenous gene Glu-1Ebx to the wild-type cultivar Bobwhite by transgenic approach. In this study, comparative seed transcriptomics and proteomics of transgenic and non-transgenic lines at different seed developmental stages were carried out to explore the changes in genes and proteins and the underlying regulatory mechanism. Results revealed that a number of genes, including genes related to SSPs, carbohydrates metabolism, amino acids metabolism, transcription, translation, and protein process were differentially enriched. Seed storage proteins displayed differential expression patterns between the transgenic and non-transgenic line, a major rise in the expression levels of gliadins were observed at 21 and 28 days post anthesis (DPA) in the transgenic line. Changes in expressions of low-molecular-weight glutenins (LMW-GSs), avenin-like proteins (ALPs), lipid transfer proteins (LTPs), and protease inhibitors (PIs) were also observed. In addition, genes related to carbohydrate metabolism were differentially expressed, which probably leads to a difference in starch component and deposition. A list of gene categories participating in the accumulation of SSPs was proposed according to the transcriptome and proteome data. Six genes from the MYB and eight genes from the NAC transcription families are likely important regulators of HMW-GSs accumulation. This study will provide data support for understanding the regulatory network of wheat storage substances. The screened candidate genes can lay a foundation for further research on the regulation mechanism of HMW-GSs.

BZIP transcription factors repress the expression of wheat (Triticum aestivum L.) high molecular weight glutenin subunit genes in vegetative tissues

High molecular weight glutenin subunits (HMW GS) represent an important fraction of endosperm-specific seed-storage proteins that provide elasticity to bread dough. Previously, the second cis-regulatory module (CRM2) was found to be one of the most conserved part of HMW GS promoters, which indicated its pre-eminent role in their gene regulation. Here, we observed that deletion of CRM2 from the promoters of the Bx7 and By8 HMW GS genes increased the leakage of their transient expression in wheat leaf tissue. The effect of a VP1, an Myb and an antisense bZIP transcription factor (TF)-binding site, potentially involved in endosperm-specific regulation within CRM2, was then studied. The activity of a Bx7 gene promoter containing a mutant CRM2 with altered VP1 and Myb TF-binding sites, but an intact bZIP TF-binding site, was similarly low to that of the wild type in leaves. Transactivation analysis and EMSA indicated the binding of TFs TabZIP34 and TabZIP115 to the Skn-1 motif GTCAT in CRM2 and the repression of Bx7 and By8 HMW GS gene promoter activity in leaves. TabZIP34 and TabZIP115 may be involved in the downregulation of HMW GS gene expression in vegetative tissues and early-stage endosperm as well its modulation during seed maturation.

Seed Storage Protein, Functional Diversity and Association with Allergy

Seed Storage Protein, Functional Diversity and Association with Allergy

The effects of germination on the composition and functional properties of hemp seed protein isolate

The effects of germination on the functional properties of hemp seed protein isolates was evaluated in this study. Significant seed storage protein degradation was observed after three days of germination, while protein surface charge was altered after one day of germination. Protein solubility increased in neutral to slightly acidic pH, with 5-day germination increasing hemp seed protein solubility from 10.3% to 21.8% at pH 7.0. Water holding capacity increased from 1.03 g H 2 O/g isolate to 1.34 g H 2 O/g isolate while oil holding capacity stayed relatively constant. Germinated protein isolates demonstrated higher foaming capacity (69.2%–111.2%) and foaming stability (53.6%–75.1%) than non-germinated protein isolates. Five-day germinated hemp seed protein isolate was the only adequate emulsifier in this study, capable of stabilizing emulsion droplets down to 2.90 μm in diameter. Hemp seed protein formed self-supporting gels at minimum of 2 or 3 w/v %, and germination increased the hardness of hemp seed protein gels (38.02 g–49.12 g for 10 w/v % gels). These findings indicate the ability to leverage germination as a clean-label process for improving the functional properties of hemp seed protein. • Hemp seed storage proteins are degraded after three days of germination. • Germinated hemp seed protein isolates are more soluble near a neutral pH. • Germination improves hemp protein WHC, foaming, emulsifying, and gelling properties. • LGC for hemp protein at pH 7 was determined at 2–3 w/v %.

Molecular peanut sensitization patterns in Lithuanian children with suspected allergic symptoms.

There are no in-depth studies describing the peanut sensitization molecular patterns in Lithuanian children. To investigate the age-related patterns of molecular peanut sensitization profiles in Lithuanian children with suspected allergic symptoms. We performed a retrospective analysis of peanut sensitization profiles in 576 Lithuanian children with possible allergic symptoms. Patient data were categorized according to age groups: 0-2, 3-6, 7-12, and 13-18 years. Specific immunoglobulin E levels to peanut molecular components: Ara h 1, Ara h 2, Ara h 3, Ara h 6, Ara h 8, Ara h 9, Ara h 15, and birch major allergen component Bet v 1 were analyzed. Sensitization to at least one peanut protein was observed in 148 (25.7%) children. In children aged ≤ 2 years, most children were sensitized to Ara h 1 - 11 (12.1%). In children aged from 3 to 6 years, the sensitization to Ara h 1 remained the most prevalent in 40 (16.6%) children. The most prevalent sensitization in children aged from 7 to 12 years was to Ara h 8 in 39 (24.5%) of them. In children aged ≥ 13 years, Ara h 8 remained the most prevalent sensitizer in 21 (24.7%) of them. One-fourth of children with suspected allergic symptoms are sensitized to at least one peanut molecular component in Lithuania. Infants and preschool children are most commonly sensitized to seed storage proteins, while school-age children to Ara h 8 allergen.

Immunological analysis of concanavalin A and canavalin in sword bean teas

Seeds of sword bean (Canavalia gladiata), a leguminous plant, is rich in proteins and cannot be eaten raw because the seed contains toxins, such as concanavalin A (ConA) and canatoxin. Therefore, the seeds are soaked, boiled, rinsed, and pounded before using them as a protein ingredient. Also, the seeds have been utilized as a natural medicine. Recently, it was reported that consuming sword bean tea alleviates allergic rhinitis symptoms. Because ConA is a known mitogen of T-cell lymphocytes, the protein may be involved in the alleviation. Therefore, we examined the detection of ConA in the extracts of sword bean teas. We prepared sword bean tea from the embryos collected after removing seed coats from the seeds immersed in water. We analyzed the proteins in the tea extract during the production process using polyacrylamide gel electrophoresis. Immunoblotting with antibodies against ConA or canavalin, a major seed storage protein, was conducted to detect the protein in the tea extract. Our result indicated that the extractable protein amount decreased after heating during the processing phase. As described above, the extracts of commercially available sword bean teas were also examined with electrophoresis and immunoblotting. The results indicated that a lower amount of ConA was detected in the tea but not canavalin.

Characterization of Seed Storage Proteins in Eight Bambara Groundnut Landraces in Burkina Faso

Characterization of Seed Storage Proteins in Eight Bambara Groundnut Landraces in Burkina Faso

Significance of Seed Storage Protein and Seed Morphological Characters in the Classification of Some Species of Malvaceae s.l.

Significance of Seed Storage Protein and Seed Morphological Characters in the Classification of Some Species of Malvaceae s.l.