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WRKYs are important regulators in plant development and stress responses. However, knowledge of this superfamily in soybean is limited. In this study, we characterized the drought- and salt-induced gene GmWRKY12 based on RNA-Seq and qRT-PCR. GmWRKY12, which is 714 bp in length, encoded 237 amino acids and grouped into WRKY II. The promoter region of GmWRKY12 included ABER4, MYB, MYC, GT-1, W-box and DPBF cis-elements, which possibly participate in abscisic acid (ABA), drought and salt stress responses. GmWRKY12 was minimally expressed in different tissues under normal conditions but highly expressed under drought and salt treatments. As a nucleus protein, GmWRKY12 was responsive to drought, salt, ABA and salicylic acid (SA) stresses. Using a transgenic hairy root assay, we further characterized the roles of GmWRKY12 in abiotic stress tolerance. Compared with control (Williams 82), overexpression of GmWRKY12 enhanced drought and salt tolerance, increased proline (Pro) content and decreased malondialdehyde (MDA) content under drought and salt treatment in transgenic soybean seedlings. These results may provide a basis to understand the functions of GmWRKY12 in abiotic stress responses in soybean.

Peptides present in foods are involved in nutritional functions by supplying amino acids; sensory functions related to taste or solubility, emulsification, etc.; and bioregulatory functions in various physiological activities. In particular, peptides have a wide range of physiological functions, including as anticancer agents and in lowering blood pressure and serum cholesterol levels, enhancing immunity, and promoting calcium absorption. Soy protein can be partially hydrolyzed enzymatically to physiologically active soy (or soybean) peptides (SPs), which not only exert physiological functions but also help amino acid absorption in the body and reduce bitterness by hydrolyzing hydrophobic amino acids from the C- or N-terminus of soy proteins. They also possess significant gel-forming, emulsifying, and foaming abilities. SPs are expected to be able to prevent and treat atherosclerosis by inhibiting the reabsorption of bile acids in the digestive system, thereby reducing blood cholesterol, low-density lipoprotein, and fat levels. In addition, soy contains blood pressure-lowering peptides that inhibit angiotensin-I converting enzyme activity and antithrombotic peptides that inhibit platelet aggregation, as well as anticancer, antioxidative, antimicrobial, immunoregulatory, opiate-like, hypocholesterolemic, and antihypertensive activities. In animal models, neuroprotective and cognitive capacity as well as cardiovascular activity have been reported. SPs also inhibit chronic kidney disease and tumor cell growth by regulating the expression of genes associated with apoptosis, inflammation, cell cycle arrest, invasion, and metastasis. Recently, various functions of soybeans, including their physiologically active functions, have been applied to health-oriented foods, functional foods, pharmaceuticals, and cosmetics. This review introduces some current results on the role of bioactive peptides found in soybeans related to health functions.

A new member of the AP2/ERF transcription factor family, GmERF3, was isolated from soybean. Sequence analysis showed that GmERF3 contained an AP2/ERF domain of 58 amino acids and two putative nuclear localization signal (NLS) domains. It belonged to a group IV protein in the ERF (ethylene response factor) subfamily as typified by a conserved N-terminal motif [MCGGAI(I/L)]. Expression of GmERF3 was induced by treatments with high salinity, drought, abscisic acid (ABA), salicylic acid (SA), jasmonic acid (JA), ethylene (ET), and soybean mosaic virus (SMV), whereas there was no significant GmERF3 mRNA accumulation under cold stress treatment. GmERF3 could bind to the GCC box and DRE/CRT element, and was targeted to the nucleus when transiently expressed in onion epidermal cells. The GmERF3 protein fused to the GAL4 DNA-binding domain to activate transcription of reporter genes in yeast. Ectopic expression of the GmERF3 gene in transgenic tobacco plants induced the expression of some PR genes and enhanced resistance against infection by Ralstonia solanacearum, Alternaria alternata, and tobacco mosaic virus (TMV), and gave tolerance to high salinity and dehydration stresses. Furthermore, overexpression of GmERF3 in transgenic tobacco led to higher levels of free proline and soluble carbohydrates compared to wild-type plants under drought conditions. The overall results suggested that GmERF3 as an AP2/ERF transcription factor may play dual roles in response to biotic and abiotic stresses in plants.

Key messageGenetic resources contributes to the sustainable protein production in soybean.Soybean is an important crop for food, oil, and forage and is the main source of edible vegetable oil and vegetable protein. It plays an important role in maintaining balanced dietary nutrients for human health. The soybean protein content is a quantitative trait mainly controlled by gene additive effects and is usually negatively correlated with agronomic traits such as the oil content and yield. The selection of soybean varieties with high protein content and high yield to secure sustainable protein production is one of the difficulties in soybean breeding. The abundant genetic variation of soybean germplasm resources is the basis for overcoming the obstacles in breeding for soybean varieties with high yield and high protein content. Soybean has been cultivated for more than 5000 years and has spread from China to other parts of the world. The rich genetic resources play an important role in promoting the sustainable production of soybean protein worldwide. In this paper, the origin and spread of soybean and the current status of soybean production are reviewed; the genetic characteristics of soybean protein and the distribution of resources are expounded based on phenotypes; the discovery of soybean seed protein-related genes as well as transcriptomic, metabolomic, and proteomic studies in soybean are elaborated; the creation and utilization of high-protein germplasm resources are introduced; and the prospect of high-protein soybean breeding is described.

An experiment was conducted to measure the effect of adding soybean oil to soybean meal (SBM) and soy protein concentrate (SPC) on apparent (AID) and standardized (SID) ileal digestibility of CP and AA by growing pigs. A second objective was to compare AID and SID of AA in a new high-protein variety of full fat soybeans (FFSB) to values obtained in other soybean products. Commercial sources of FFSB (FFSB-CV), SBM, and SPC, and of a new high-protein variety of FFSB (FFSB-HP) were used in the experiment. Four diets were prepared using each soybean product as the sole source of CP and AA in 1 diet. Two additional diets were formulated by adding soybean oil (7.55 and 7.35%, respectively) to the diets containing SBM and SPC. A nitrogen-free diet was also used to measure basal endogenous losses of CP and AA. The 2 sources of FFSB were extruded at 150 degrees C before being used in the experiment. Seven growing barrows (initial BW = 26.2 kg) were prepared with a T-cannula in the distal ileum and allotted to a 7 x 7 Latin square design. Ileal digesta were collected from the pigs on d 6 and 7 of each period. All digesta samples were lyophilized and analyzed for DM, CP, AA, and chromium, and values for AID and SID of CP and AA were calculated. The addition of oil improved (P < 0.05) the SID of most indispensable AA in SBM and SPC. The SID for 6 of the indispensable AA in FFSB-HP were greater (P < 0.05) than in FFSB-CV, and the SID for all indispensable AA except Met was greater (P < 0.05) in FFSB-HP than in SBM. However, the SID for most AA in FFSB-HP was similar to SBM with oil and SPC, but these values were lower (P < 0.05) than in SPC with oil. In conclusion, the addition of oil improved the SID of most AA in SBM and SPC fed to growing pigs, and the SID of AA in FFSB-HP were greater than in SBM and similar to the SID of AA in SBM with oil and in SPC.

The content of protein, moisture content and essential amino acids in conventional and genetically modified soybean grain and selected soybean products (soybean pâté, soybean drink, soybean dessert, tofu) was analyzed in this paper. The following comparative analysis of these products has not yet been carried out. No differences were observed in the amino acid profiles of soybeans and soybean products. The presence of essential amino acids was confirmed except for tryptophan. Its absence, however, may be due not to its absence in the raw material, but to its decomposition as a result of the acid hydrolysis of the sample occurring during its preparation for amino acid determination. Regardless of the type of soybean grain, the content of protein, moisture content and essential amino acids was similar (statistically insignificant difference). Thus, the type of raw material did not determine these parameters. There was a significant imbalance in the quantitative composition of essential amino acids in individual soybean products. Only statistically significant variation was found in genetically modified and conventional soybean pâté. Moreover, in each soy product their amount was lower irrespective of the raw material from which they were manufactured. Therefore, the authors indicate the necessity of enriching soybean products with complete protein to increase their nutritional value.

An experiment was conducted to determine the apparent (AID) and standardized (SID) ileal digestibility of CP and AA in weanling pigs of 4 soybean products, fish meal, and casein. The 4 soybean products were conventional dehulled soybean meal (SBM), soy protein isolate (SPI), fermented soybean meal (FSBM), and enzyme-treated soybean meal (ESBM). Seven weanling barrows (initial BW: 10.9 ± 2.3 kg) were individually fitted with a T-cannula in the distal ileum. The barrows were allotted to a 7 x 7 Latin square design with 7 diets and seven 7-d periods. Six cornstarch-based diets were prepared using each of the protein sources as the sole source of CP and AA. An N-free diet was used to measure basal endogenous losses of CP and AA. Results showed that except for Lys, the AID and SID of AA in FSBM was not different from SBM, and with a few exceptions, the AID and SID of most AA in SBM, FSBM, and ESBM were not different from each other and from the AID and SID of AA in fish meal. Likewise, the AID and SID of AA in ESBM and SPI were not different, but the AID and SID of most AA in SPI were greater (P < 0.05) than in SBM and FSBM. The AID and SID of most AA in SPI were not different from the AID and SID of AA in casein. In conclusion, FSBM and ESBM had similar SID of most AA as SBM, but SPI has the greatest SID of AA among the 4 soybean products. Casein had the greatest SID of AA among the protein sources studied.

Limosilactobacillus reuteri DSM17938 is one of the most pivotal probiotics, whose general beneficial effects on the intestinal microbiota are well recognized. Enhancing their growth and metabolic activity can effectively regulate the equilibrium of intestinal microbiota, leading to improved physical health. A common method to promote the growth of Lactobacillus is the addition of prebiotics. Current research suggests that proteins and their hydrolysates from different sources with potential prebiotic activity can also promote the growth of probiotics. In this study, soybean proteins and peptides were effective in promoting the growth, organic acid secretion, and adhesive properties of Limosilactobacillus reuteri DSM17938 to Caco-2 cells. These results illustrate the feasibility of soybean proteins and peptides as prebiotics, providing theoretical and practical advantages for their application.

Soybean meal (SBM) is extensively used as a predominant protein source in animal feed. However, raw soybean cannot be directly utilized in animal feed, due to the presence of the Kunitz trypsin inhibitor (KTi) and the Bowman–Birk protease inhibitor (BBi). These antinutritional factors inhibit the digestive enzymes in animals, trypsin and chymotrypsin, resulting in poor animal performance. To inactivate the activity of protease inhibitors, SBM is subjected to heat processing, a procedure that can negatively impact the soybean protein quality. Thus, it would be beneficial to develop soybean varieties with little or no trypsin inhibitors. In this study, we report on the creation of experimental soybean lines with significantly reduced levels of Bowman–Birk protease inhibitors. RNA interference (RNAi) technology was employed to generate several transgenic soybean lines. Some of these BBi knockdown soybean lines showed significantly lower amounts of both trypsin and chymotrypsin inhibitor activities. Western blot analysis revealed the complete absence of BBi in selected RNAi-derived lines. RNA sequencing (RNAseq) analysis demonstrated a drastic reduction in the seed-specific expression of BBi genes in the transgenic soybean lines during seed development. Confocal fluorescence immunolabeling studies showed that the accumulation of BBi was drastically diminished in BBi knockdown lines compared to wild-type soybeans. The absence of BBi in the transgenic soybean did not alter the overall protein, oil, and sulfur amino acid content of the seeds compared to wild-type soybeans. The seed protein from the BBi knockdown lines were more rapidly hydrolyzed by trypsin and chymotrypsin compared to the wild type, indicating that the absence of BBi enhances protein digestibility. Our study suggests that these BBi knockdown lines could be a valuable resource in order for plant breeders to incorporate this trait into commercial soybean cultivars, potentially enabling the use of raw soybeans in animal feed.

Key messageProteomic and lipidomics analyses of WT and GmDGAT1-2 transgenic soybeans showed that GmDGAT1-2 over-expression induced lipoxygenase down-regulatation and oleoin up-regulatation, which significantly changed the compositions and total fatty acid.The main goal of soybean breeding is to increase the oil content. Diacylglycerol acyltransferase (DGAT) is a key rate-limiting enzyme in fatty acid metabolism and may regulate oil content. Herein, 10 GmDGAT genes were isolated from soybean and transferred into wild-type (WT) Arabidopsis. The total fatty acid was 1.2 times higher in T3GmDGAT1-2 transgenic Arabidopsis seeds than in WT. Therefore, GmDGAT1-2 was transferred into WT soybean (JACK), and four T3 transgenic soybean lines were obtained. The results of high-performance gas chromatography and Soxhlet extractor showed that, compared with those of JACK, oleic acid (18:1), and total fatty acid levels in transgenic soybean plants were much higher, but linoleic acid (18:2) was lower than WT. Palmitic acid (16:0), stearic acid (18:0), and linolenic acid (18:3) were not significantly different. For mechanistic studies, 436 differentially expressed proteins (DEPs) and 180 differentially expressed metabolites (DEMs) were identified between WT (JACK) and transgenic soybean pods using proteomic and lipidomics analyses. Four lipoxygenase proteins were down-regulated in linoleic acid metabolism while four oleosin proteins were up-regulated in the final oil formation. The results showed an increase in the total fatty acid and 18:1 composition, and a decrease in the 18:2 composition of fatty acid. Our study brings new insights into soybean genetic transformation and the deep study of molecular mechanism that changes the total fatty acid, 18:1, and 18:2 compositions in GmDGAT1-2 transgenic soybean.