Identification and functional analysis of GmPasL regulating pod color in vegetable soybean.

BACKGROUND: Vegetable soybean is rich in nutrients and has a unique flavor. It is highly preferred by people because of its pharmacological activities, including those that regulate the intestines and lower blood pressure. The pod color of vegetable soybeans is an important quality that indicates their freshness and has a significant impact on their commercialization. RESULTS: In this study, pod color was evaluated in 301 vegetable soybean accessions collected from various regions. Genome-wide association analysis was carried out using the Mixed linear model (MLM), a total of 18 quantitative trait loci including 117 SNPs were detected. Two significant QTLs located on chromosomes 6 (qGPCL4 /qGPCa1/qGPCb2) and 18 (qGPCL10/qGPCb3) were consistently detected across different variables. Based on gene functional annotation, 30 candidate genes were identified in these two candidate intervals. Combined with transcriptome analysis, Glyma.18g241700 has been identified as a candidate gene for regulating pod color in vegetable soybeans. Glyma.18g241700 encodes a chlorophyll photosystem I subunit XI. which localizes to the chloroplast named GmPsaL, qRT-PCR analysis showed that GmPsaL was specifically highly expressed in developing pods. Furthermore, overexpression of GmPsaL in transgenetic Arabidopsis plants produced dark green pods. CONCLUSIONS: These findings may be useful for clarifying the genetic basis of the pod color of vegetable soybeans. The identified candidate genes may be useful for the genetic improvement of the appearance quality of vegetable soybeans.

Expansion and diversification of the Glycine max (Gm) ERD15-like subfamily of the PAM2-like superfamily.

MAIN CONCLUSION: Despite modulating senescence and drought responses, the GmERD15-like subfamily members are differentially induced by multiple stresses and diverge partially in stress signaling functions. The PAM2 motif represents a binding site for poly (A)-binding proteins (PABPs), often associated with RNA metabolism regulation. The PAM2-containing protein ERD15 stands out as a critical regulator of diverse stress responses in plants. Despite the relevance of the PAM2 motif, a comprehensive analysis of the PAM2 superfamily and ERD15-like subfamily in the plant kingdom is lacking. Here, we provide an extensive in silico analysis of the PAM2 superfamily and the ERD15-like subfamily in soybean, using Arabidopsis and rice sequences as prototypes. The Glycine max ERD15-like subfamily members were clustered in pairs, likely originating from DNA-based gene duplication, as the paralogs display high sequence conservation, similar exon/intron genome organization, and are undergoing purifying selection. Complementation analyses of an aterd15 mutant demonstrated that the plant ERD15-like subfamily members are functionally redundant in response to drought, osmotic stress, and dark-induced senescence. Nevertheless, the soybean members displayed differential expression profiles, biochemical activity, and subcellular localization, consistent with functional diversification. The expression profiles of Glyma04G138600 under salicylic acid (SA) and abscisic acid (ABA) treatments differed oppositely from those of the other GmERD15-like genes. Abiotic stress-induced coexpression analysis with soybean PABPs showed that Glyma04G138600 was clustered separately from other GmERD15s. In contrast to the AtERD15 stress-induced nuclear redistribution, Glyma04G138600 and Glyma02G260800 localized to the cytoplasm, while Glyma03G131900 fractionated between the cytoplasm and nucleus under normal and stress conditions. These data collectively indicate that despite modulating senescence and drought responses, the GmERD15-like subfamily members are differentially induced by multiple stresses and may diverge partially in stress signaling functions.

Deciphering the genetic basis of salinity tolerance in a diverse panel of cultivated and wild soybean accessions by genome-wide association mapping.

KEY MESSAGE: In a genome-wide association study involving 269 cultivated and wild soybean accessions, potential salt tolerance donors were identified along with significant markers and candidate genes, such as GmKUP6 and GmWRKY33. Salt stress remains a significant challenge in agricultural systems, notably impacting soybean productivity worldwide. A comprehensive genome-wide association study (GWAS) was conducted to elucidate the genetic underpinnings of salt tolerance and identify novel source of salt tolerance among soybean genotypes. A diverse panel comprising 269 wild and cultivated soybean accessions was subjected to saline stress under controlled greenhouse conditions. Phenotypic data revealed that salt tolerance of soybean germplasm accessions was heavily compromised by the accumulation of sodium and chloride, as indicated by highly significant positive correlations of leaf scorching score with leaf sodium/chloride content. The GWAS analysis, leveraging a dataset of 32,832 SNPs, unveiled 32 significant marker-trait associations (MTAs) across seven traits associated with salt tolerance. These markers explained a substantial portion of the phenotypic variation, ranging from 14 to 52%. Notably, 11 markers surpassed Bonferroni's correction threshold, exhibiting highly significant associations with the respective traits. Gene Ontology enrichment analysis conducted within a 100 Kb range of the identified MTAs highlighted candidate genes such as potassium transporter 6 (GmKUP6), cation hydrogen exchanger (GmCHX15), and GmWRKY33. Expression levels of GmKUP6 and GmWRKY33 significantly varied between salt-tolerant and salt-susceptible soybean accessions under salt stress. The genetic markers and candidate genes identified in this study hold promise for developing soybean varieties resilient to salinity stress, thereby mitigating its adverse effects.

Genetic transformation of GmFBX322 gene and salt tolerance physiology in soybean.

Soybean is one of the most important food crops, breeding salt-tolerant soybean varieties is of great significance to alleviate soybean shortage. In this study, the F-box protein family homologous gene GmFBX322 was cloned from the soybean variety Williams 82 and overexpressed in the Shennong 9 soybean variety to further study and explore the physiological mechanism of soybean salt tolerance. GmFBX322 was constructed on the vector pTF101:35S, and integrated into the genome of Shennong 9 soybean variety by Agrobacterium EHA101-mediated cotyledonary node transformation technology, and 4 overexpressed transgenic lines were obtained, molecular assays were performed on the transformed plants. The expression of GmFBX322 was detected by qRT-PCR and it was found that the leaves of the 4 transgenic lines increased by 2.49, 2.46, 2.77, 2.95 times compared with the wild type; after salt treatment for 12 hours, it was found that the expression of wild type Shennong 9 Inducible expression of GmFBX322. After 72 hours of salt treatment, the leaves of wild-type Shennong 9 soybean plants showed obvious wilting and chlorosis, while the leaves of GmFBX322 plants overexpressing GmFBX322 showed no obvious changes. The leaves were taken at 0, 6, 12, 24, and 48 hours of salt stress to determine the antioxidant activity. Ability and osmotic adjustment level, etc. The results showed that the catalase activity in the leaves of the transgenic lines 2265, 2267, 2269, and 2271 was 2.47, 2.53, 3.59, 2.96 times that of the wild-type plant after 48 hours of salt treatment; the soluble sugar content was 1.22, 1.14, and 1.22 of the wild-type plant. 1.14, 1.57 times; the proline content is 2.20, 1.83, 1.65, 1.84 times of the wild type. After comparing the physiological indicators determined by the experiment, the transgenic lines performed better than the control group, indicating that overexpression of GmFBX322 can enhance the salt tolerance of soybean plants. To verify the function of GmFBX322 gene related to stress resistance, add it to the candidate gene of stress resistance, and provide scientific basis for the selection and breeding of salt-tolerant varieties.

How do drought and heat affect the response of soybean seed yield to elevated O3? An analysis of 15 seasons of free-air O3 concentration enrichment studies.

The coincidence of rising ozone concentrations ([O3]), increasing global temperatures, and drought episodes is expected to become more intense and frequent in the future. A better understanding of the responses of crop yield to elevated [O3] under different levels of drought and high temperature stress is, therefore, critical for projecting future food production potential. Using a 15-year open-air field experiment in central Illinois, we assessed the impacts of elevated [O3] coupled with variation in growing season temperature and water availability on soybean seed yield. Thirteen soybean cultivars were exposed to a wide range of season-long elevated [O3] in the field using free-air O3 concentration enrichment. Elevated [O3] treatments reduced soybean seed yield from as little as 5.3% in 2005 to 35.2% in 2010. Although cultivars differed in yield response to elevated [O3] (R), ranging from 17.5% to -76.4%, there was a significant negative correlation between R and O3 dosage. Soybean cultivars showed greater seed yield losses to elevated [O3] when grown at drier or hotter conditions compared to wetter or cooler years, because the hotter and drier conditions were associated with greater O3 treatment. However, year-to-year variation in weather conditions did not influence the sensitivity of soybean seed yield to a given increase in [O3]. Collectively, this study quantitatively demonstrates that, although drought conditions or warmer temperatures led to greater O3 treatment concentrations and O3-induced seed yield reduction, drought and temperature stress did not alter soybean's sensitivity to O3. Our results have important implications for modeling the effects of rising O3 pollution on crops and suggest that altering irrigation practices to mitigate O3 stress may not be effective in reducing crop sensitivity to O3.

Genome-wide association study provides new insight into the underlying mechanism of drought tolerance during seed germination stage in soybean.

Drought is one of the major environmental issues that reduce crop yield. Seed germination is a crucial stage of plant development in all crop plants, including soybean. In soybean breeding, information about genetic mechanism of drought tolerance has great importance. However, at germination stage, there is relatively little knowledge on the genetic basis of soybean drought resistance. The objective of this work was to find the quantitative trait nucleotides (QTNs) linked to drought tolerance related three traits using a genome-wide association study (GWAS), viz., germination rate (GR), root length (RL), and whole seedling length (WSL), using germplasm population of 240 soybean PIs with 34,817 SNPs genotype data having MAF > 0.05. It was observed that heritability (H2) for GR, WSL, and RL across both environments (2020, and 2019) were high in the range of 0.76-0.99, showing that genetic factors play a vital role in drought tolerance as compared to environmental factors. A number of 23 and 27 QTNs were found to be linked to three traits using MLM and mrMLM, respectively. Three significant QTNs, qGR8-1, qWSL13-1, and qRL-8, were identified using both MLM and mrMLM methods among these QTNs. QTN8, located on chromosome 8 was consistently linked to two traits (GR and RL). The area (± 100 Kb) associated with this QTN was screened for drought tolerance based on gene annotation. Fifteen candidate genes were found by this screening. Based on the expression data, four candidate genes i.e. Glyma08g156800, Glyma08g160000, Glyma08g162700, and Glyma13g249600 were found to be linked to drought tolerance regulation in soybean. Hence, the current study provides evidence to understand the genetic constitution of drought tolerance during the germination stage and identified QTNs or genes could be utilized in molecular breeding to enhance the yield under drought stress.

The rhizobacterial Priestia megaterium strain SH-19 mitigates the hazardous effects of heat stress via an endogenous secondary metabolite elucidation network and molecular regulation signalling.

Global warming is a leading environmental stress that reduces plant productivity worldwide. Several beneficial microorganisms reduce stress; however, the mechanism by which plant-microbe interactions occur and reduce stress remains to be fully elucidated. The aim of the present study was to elucidate the mutualistic interaction between the plant growth-promoting rhizobacterial strain SH-19 and soybeans of the Pungsannamul variety. The results showed that SH-19 possessed several plant growth-promoting traits, such as the production of indole-3-acetic acid, siderophore, and exopolysaccharide, and had the capacity for phosphate solubilisation. The heat tolerance assay showed that SH-19 could withstand temperatures up to 45 °C. The strain SH-19 was identified as P. megaterium using the 16S ribosomal DNA gene sequence technique. Inoculation of soybeans with SH-19 improved seedling characteristics under high-temperature stress. This may be due to an increase in the endogenous salicylic acid level and a decrease in the abscisic acid level compared with the negative control group. The strain of SH-19 increased the activity of the endogenous antioxidant defense system, resulting in the upregulation of GSH (44.8%), SOD (23.1%), APX (11%), and CAT (52.6%). Furthermore, this study involved the transcription factors GmHSP, GmbZIP1, and GmNCED3. The findings showed upregulation of the two transcription factors GmbZIP1 (17%), GmNCED3 (15%) involved in ABA biosynthesis and induced stomatal regulation, similarly, a downregulation of the expression pattern of GmHSP by 25% was observed. Overall, the results of this study indicate that the strain SH-19 promotes plant growth, reduces high-temperature stress, and improves physiological parameters by regulating endogenous phytohormones, the antioxidant defense system, and genetic expression. The isolated strain (SH-19) could be commercialized as a biofertilizer.

Exploration and genetic analyses of canopy leaf pigmentation changes in soybean (Glycine max L.): unveiling a novel phenotype.

KEY MESSAGE: Pigmentation changes in canopy leaves were first reported, and subsequent genetic analyses identified a major QTL associated with levels of pigmentation changes, suggesting Glyma.06G202300 as a candidate gene. An unexpected reddish-purple pigmentation in upper canopy leaves was discovered during the late reproductive stages in soybean (Glycine max L.) genotypes. Two sensitive genotypes, 'Uram' and PI 96983, exhibited anomalous canopy leaf pigmentation changes (CLPC), while 'Daepung' did not. The objectives of this study were to: (i) characterize the physiological features of pigmented canopy leaves compared with non-pigmented leaves, (ii) evaluate phenotypic variation in a combined recombinant inbred line (RIL) population (N = 169 RILs) under field conditions, and (iii) genetically identify quantitative trait loci (QTL) for CLPC via joint population linkage analysis. Comparison between pigmented and normal leaves revealed different Fv/Fm of photosystem II, hyperspectral reflectance, and cellular properties, suggesting the pigmentation changes occur in response to an undefined abiotic stress. A highly significant QTL was identified on chromosome 6, explaining ~ 62.8% of phenotypic variance. Based on the QTL result, Glyma.06G202300 encoding flavonoid 3'-hydroxylase (F3'H) was identified as a candidate gene. In both Uram and PI 96983, a 1-bp deletion was confirmed in the third exon of Glyma.06G202300 that results in a premature stop codon in both Uram and PI 96983 and a truncated F3'H protein lacking important domains. Additionally, gene expression analyses uncovered significant differences between pigmented and non-pigmented leaves. This is the first report of a novel symptom and an associated major QTL. These results will provide soybean geneticists and breeders with valuable knowledge regarding physiological changes that may affect soybean production. Further studies are required to elucidate the causal environmental stress and the underlying molecular mechanisms.

Mechanisms of Lanthanum-mediated mitigation of salt stress in soybean (Glycine max L.).

Salinity is considered one of the abiotic stresses that have the greatest impact on soybean production worldwide. Lanthanum (La) is a rare earth element that can reduce adverse conditions on plant growth and productivity. However, the regulatory mechanism of La-mediated plant response to salt stress has been poorly studied, particularly in soybeans. Therefore, our study investigated the mechanisms of La-mediated salt stress alleviation from the perspectives of the antioxidant system, subcellular structure, and metabolomics responses. The results indicated that salt stress altered plant morphology and biomass, resulting in an increase in peroxidation, inhibition of photosynthesis, and damage to leaf structure. Exogenous La application effectively promoted the activity of superoxide dismutase (SOD) and peroxidase (POD), as well as the soluble protein content, while decreasing the Na+ content and Na+/K+ ratio in roots and leaves, and reducing oxidative damage. Moreover, transmission electron microscopy (TEM) demonstrated that La prevented the disintegration of chloroplasts. Fourier-transform infrared spectroscopy (FTIR) analysis further confirmed that La addition mitigated the decline in protein, carbohydrates, and pectin levels in the leaves. Lanthanum decreased the leaf flavonoid content and synthesis by inhibiting the content of key substances in the phenylalanine metabolism pathway during NaCl exposure. Collectively, our research indicates that La reduces cell damage by regulating the antioxidant system and secondary metabolite synthesis, which are important mechanisms for the adaptive response of soybean leaves, thereby improving the salt tolerance of soybeans.

Soybean ethylene response factors GmENS1 and GmENS2 promote nodule senescence.

The final phase in root nodule development is nodule senescence. The mechanism underlying the initiation of nodule senescence requires further elucidation. In this study, we investigate the intrinsic signals governing soybean (Glycine max L. Merr.) nodule senescence, uncovering ethylene as a key signal in this intricate mechanism. Two AP2/ethylene response factor (ERF) transcription factor (TF) genes, GmENS1 and GmENS2 (Ethylene-responsive transcription factors required for Nodule Senescence), exhibit heightened expression levels in both aged nodules and nodules treated with ethylene. An overexpression of either GmENS1 or GmENS2 accelerates senescence in soybean nodules, whereas the knockout or knockdown of both genes delays senescence and enhances nitrogenase activity. Furthermore, our findings indicate that GmENS1 and GmENS2 directly bind to the promoters of GmNAC039, GmNAC018, and GmNAC030, encoding 3 NAC (NAM, ATAF1/2, and CUC2) TFs essential for activating soybean nodule senescence. Notably, the nodule senescence process mediated by GmENS1 or GmENS2 overexpression is suppressed in the soybean nac039/018/030 triple mutant compared with the wild-type control. These data indicate GmENS1 and GmENS2 as pivotal TFs mediating ethylene-induced nodule senescence through the direct activation of GmNAC039/GmNAC018/GmNAC030 expression in soybean.

Bacillus suppresses nitrogen efficiency of soybean-rhizobium symbiosis through regulation of nitrogen-related transcriptional and microbial patterns.

The regulation of legume-rhizobia symbiosis by microorganisms has obtained considerable interest in recent research, particularly in the common rhizobacteria Bacillus. However, few studies have provided detailed explanations regarding the regulatory mechanisms involved. Here, we investigated the effects of Bacillus (Bac.B) on Bradyrhizobium-soybean (Glycine max) symbiosis and elucidated the underlying ecological mechanisms. We found that two Bradyrhizobium strains (i.e. Bra.Q2 and Bra.D) isolated from nodules significantly promoted nitrogen (N) efficiency of soybean via facilitating nodule formation, thereby enhanced plant growth and yield. However, the intrusion of Bac.B caused a reverse shift in the synergistic efficiency of N2 fixation in the soybean-Bradyrhizobium symbiosis. Biofilm formation and naringenin may be importantin suppression of Bra.Q2 growth regulated by Bac.B. In addition, transcriptome and microbiome analyses revealed that Bra.Q2 and Bac.B might interact to regulateN transport and assimilation, thus influence the bacterial composition related to plant N nutrition in nodules. Also, the metabolisms of secondary metabolites and hormones associated with plant-microbe interaction and growth regulation were modulated by Bra.Q2 and Bac.B coinoculation. Collectively, we demonstrate that Bacillus negatively affects Bradyrhizobium-soybean symbiosis and modulate microbial interactions in the nodule. Our findings highlight a novel Bacillus-based regulation to improve N efficiency and sustainable agricultural development.

GmANKTM21 Positively Regulates Drought Tolerance and Enhanced Stomatal Response through the MAPK Signaling Pathway in Soybean.

Drought stress is one of the significant abiotic stresses that limit soybean (Glycine max [L.] Merr.) growth and production. Ankyrin repeat (ANK) proteins, being highly conserved, occupy a pivotal role in diverse biological processes. ANK genes were classified into nine subfamilies according to conserved domains in the soybean genome. However, the function of ANK-TM subfamily proteins (Ankyrin repeat proteins with a transmembrane domain) in the abiotic-stress response to soybean remains poorly understood. In this study, we first demonstrated the subcellular localization of GmANKTM21 in the cell membrane and nucleus. Drought stress-induced mRNA levels of GmANKTM21, which encodes proteins belonging to the ANK-TM subfamily, Transgenic 35S:GmANKTM21 soybean improved drought tolerance at the germination and seedling stages, with higher stomatal closure in soybean, lower water loss, lower malondialdehyde (MDA) content, and less reactive oxygen species (ROS) production compared with the wild-type soybean (Dongnong50). RNA-sequencing (RNA-seq) and RT-qPCR analysis of differentially expressed transcripts in overexpression of GmANKTM21 further identified potential downstream genes, including GmSPK2, GmSPK4, and GmCYP707A1, which showed higher expression in transgenic soybean, than those in wild-type soybean and KEGG enrichment analysis showed that MAPK signaling pathways were mostly enriched in GmANKTM21 overexpressing soybean plants under drought stress conditions. Therefore, we demonstrate that GmANKTM21 plays an important role in tolerance to drought stress in soybeans.

Resilience of soybean genotypes to drought stress during the early vegetative stage.

Drought stress poses a significant risk to soybean production, as it relies on optimum rainfall under rainfed conditions. Exposure to brief dry periods during early vegetative growth impacts soybean growth and development. Choosing a genotype that can withstand stress with minimal impact on physiology and growth might help sustain biomass or yields under low rainfall conditions. Therefore, this study characterized 64 soybean genotypes for traits associated with drought tolerance during the early vegetative stage under two soil moisture treatments, 100% evapotranspiration (well-watered) and 50% evapotranspiration (drought), using the Soil-Plant-Atmosphere Research (SPAR) units. Eighteen morpho-physiological traits responses were assessed, and their relationship with the early vegetative drought tolerance was investigated. Drought stress significantly increased root weight, root volume, and root-to-shoot ratio but reduced shoot weight. Drought-stressed plants increased the canopy temperature by 3.1 °C. Shoot weight positively correlated with root surface area (r = 0.52, P < 0.001) and root weight (r = 0.65, P < 0.001). There was a strong negative correlation between shoot weight and root-to-shoot ratio (P < 0.01). Further, the combined drought response index was strongly associated with the root response index and weakly with the physiological response index. These findings suggest that genotypes (S55-Q3 and R2C4775) with high above-ground biomass with a balanced root-to-shoot ratio improves drought tolerance during the early vegetative. These genotypes could serve as valuable genetic resources to dissect the molecular networks underlying drought tolerance and ultimately be used in breeding programs to improve root ability at the early vegetative stage.

Natural variation of domestication-related genes contributed to latitudinal expansion and adaptation in soybean.

Soybean is a major source of protein and edible oil worldwide. Originating from the Huang-Huai-Hai region, which has a temperate climate, soybean has adapted to a wide latitudinal gradient across China. However, the genetic mechanisms responsible for the widespread latitudinal adaptation in soybean, as well as the genetic basis, adaptive differentiation, and evolutionary implications of theses natural alleles, are currently lacking in comprehensive understanding. In this study, we examined the genetic variations of fourteen major gene loci controlling flowering and maturity in 103 wild species, 1048 landraces, and 1747 cultivated species. We found that E1, E3, FT2a, J, Tof11, Tof16, and Tof18 were favoured during soybean improvement and selection, which explained 75.5% of the flowering time phenotypic variation. These genetic variation was significantly associated with differences in latitude via the LFMM algorithm. Haplotype network and geographic distribution analysis suggested that gene combinations were associated with flowering time diversity contributed to the expansion of soybean, with more HapA clustering together when soybean moved to latitudes beyond 35°N. The geographical evolution model was developed to accurately predict the suitable planting zone for soybean varieties. Collectively, by integrating knowledge from genomics and haplotype classification, it was revealed that distinct gene combinations improve the adaptation of cultivated soybeans to different latitudes. This study provides insight into the genetic basis underlying the environmental adaptation of soybean accessions, which could contribute to a better understanding of the domestication history of soybean and facilitate soybean climate-smart molecular breeding for various environments.

A new biostimulant derived from soybean by-products enhances plant tolerance to abiotic stress triggered by ozone.

BACKGROUND: Tropospheric ozone is an air pollutant that causes negative effects on vegetation, leading to significant losses in crop productivity. It is generated by chemical reactions in the presence of sunlight between primary pollutants resulting from human activity, such as nitrogen oxides and volatile organic compounds. Due to the constantly increasing emission of ozone precursors, together with the influence of a warming climate on ozone levels, crop losses may be aggravated in the future. Therefore, the search for solutions to mitigate these losses becomes a priority. Ozone-induced abiotic stress is mainly due to reactive oxygen species generated by the spontaneous decomposition of ozone once it reaches the apoplast. In this regard, compounds with antioxidant activity offer a viable option to alleviate ozone-induced damage. Using enzymatic technology, we have developed a process that enables the production of an extract with biostimulant properties from okara, an industrial soybean byproduct. The biostimulant, named as OEE (Okara Enzymatic Extract), is water-soluble and is enriched in bioactive compounds present in okara, such as isoflavones. Additionally, it contains a significant fraction of protein hydrolysates contributing to its functional effect. Given its antioxidant capacity, we aimed to investigate whether OEE could alleviate ozone-induced damage in plants. For that, pepper plants (Capsicum annuum) exposed to ozone were treated with a foliar application of OEE. RESULTS: OEE mitigated ozone-induced damage, as evidenced by the net photosynthetic rate, electron transport rate, effective quantum yield of PSII, and delayed fluorescence. This protection was confirmed by the level of expression of genes associated with photosystem II. The beneficial effect was primarily due to its antioxidant activity, as evidenced by the lipid peroxidation rate measured through malondialdehyde content. Additionally, OEE triggered a mild oxidative response, indicated by increased activities of antioxidant enzymes in leaves (catalase, superoxide dismutase, and guaiacol peroxidase) and the oxidative stress index, providing further protection against ozone-induced stress. CONCLUSIONS: The present results support that OEE protects plants from ozone exposure. Taking into consideration that the promotion of plant resistance against abiotic damage is an important goal of biostimulants, we assume that its use as a new biostimulant could be considered.

Enhancing saline stress tolerance in soybean seedlings through optimal NH4+/NO3- ratios: a coordinated regulation of ions, hormones, and antioxidant potential.

BACKGROUND: Nitrogen (N) availability is crucial in regulating plants' abiotic stress resistance, particularly at the seedling stage. Nevertheless, plant responses to N under salinity conditions may vary depending on the soil's NH4+ to NO3- ratio. METHODS: In this study, we investigated the effects of different NH4+:NO3- ratios (100/0, 0/100, 25/75, 50/50, and 75/25) on the growth and physio-biochemical responses of soybean seedlings grown under controlled and saline stress conditions (0-, 50-, and 100-mM L- 1 NaCl and Na2SO4, at a 1:1 molar ratio). RESULTS: We observed that shoot length, root length, and leaf-stem-root dry weight decreased significantly with increased saline stress levels compared to control. Moreover, there was a significant accumulation of Na+, Cl-, hydrogen peroxide (H2O2), and malondialdehyde (MDA) but impaired ascorbate-glutathione pools (AsA-GSH). They also displayed lower photosynthetic pigments (chlorophyll-a and chlorophyll-b), K+ ion, K+/Na+ ratio, and weakened O2•--H2O2-scavenging enzymes such as superoxide dismutase, catalase, peroxidase, monodehydroascorbate reductase, glutathione reductase under both saline stress levels, while reduced ascorbate peroxidase, and dehydroascorbate reductase under 100-mM stress, demonstrating their sensitivity to a saline environment. Moreover, the concentrations of proline, glycine betaine, total phenolic, flavonoids, and abscisic acid increased under both stresses compared to the control. They also exhibited lower indole acetic acid, gibberellic acid, cytokinins, and zeatine riboside, which may account for their reduced biomass. However, NH4+:NO3- ratios caused a differential response to alleviate saline stress toxicity. Soybean seedlings supplemented with optimal ratios of NH4+:NO3- (T3 = 25:75 and T = 4 50:50) displayed lower Na+ and Cl- and ABA but improved K+ and K+/Na+, pigments, growth hormones, and biomass compared to higher NH4+:NO3- ratios. They also exhibited higher O2•--H2O2-scavenging enzymes and optimized H2O2, MDA, and AsA-GSH pools status in favor of the higher biomass of seedlings. CONCLUSIONS: In summary, the NH4+ and NO3- ratios followed the order of 50:50 > 25:75 > 0:100 > 75:25 > 100:0 for regulating the morpho-physio-biochemical responses in seedlings under SS conditions. Accordingly, we suggest that applying optimal ratios of NH4+ and NO3- (25/75 and 50:50) can improve the resistance of soybean seedlings grown in saline conditions.

CRISPR-Cas9-mediated editing of GmARM improves resistance to multiple stresses in soybean.

The growth and development of soybean plants can be affected by both abiotic and biotic stressors, such as saline-alkali stress and Phytophthora root rot. In this study, we identified a stress-related gene-GmARM-whose promoter contained several hormone-response and stress-regulatory elements, including ABRE, TCA element, STRE, and MBS. qRT-PCR analysis showed that the expression of GmARM was the highest in seeds at 55 days after flowering. Furthermore, this gene was upregulated after exposure to saline-alkali stress and Phytophthora root rot infection at the seedling stage. Thus, we generated GmARM mutants using the CRISPR-Cas9 system to understand the role of this gene in stress response. T3 plants showed significantly improved salt tolerance, alkali resistance, and disease resistance, with a significantly higher survival rate than the wildtype plants. Moreover, mutations in GmARM affected the expression of related stress-resistance genes, indicating that GmARM mutants achieved multiple stress tolerance. Therefore, this study provides a foundation for further exploration of the genes involved in resistance to multiple stresses in soybean that can be used for breeding multiple stress-resistance soybean varieties.

GmHXK2 promotes the salt tolerance of soybean seedlings by mediating AsA synthesis, and auxin synthesis and distribution.

BACKGROUND: Salt is an important factor that affects crop productivity. Plant hexokinases (HXKs) are key enzymes in the glycolytic pathway and sugar signaling transduction pathways of plants. In previous studies, we identified and confirmed the roles of GmHXK2 in salt tolerance. RESULTS: In this study, we analyzed the tissue-specific expression of GmHXK2 at different growth stages throughout the plant's life cycle. The results showed that GmHXK2 was expressed significantly in all tissues at vegetative stages, including germination and seedling. However, no expression was detected in the pods, and there was little expression in flowers during the later mature period. Arabidopsis plants overexpressing the GmHXK2 (OE) had more lateral roots. The OE seedlings also produced higher levels of auxin and ascorbic acid (AsA). Additionally, the expression levels of genes PMM, YUC4/YUC6/YUC8, and PIN/LAX1,LAX3, which are involved respectively in the synthesis of AsA and auxin, as well as polar auxin transport, were upregulated in OE plants. This upregulation occurred specifically under exogenous glucose treatment. AtHKT1, AtSOS1, and AtNHX1 were up-regulated in OE plants under salt stress, suggesting that GmHXK2 may modulate salt tolerance by maintaining ion balance within the cells and alleviating damage caused by salt stress. Additionally, we further confirmed the interaction between GmHXK2 and the protein GmPMM through yeast two-hybridization and bimolecular fluorescence complementation assays, respectively. CONCLUSION: The expression of GmHXK2 gene in plants is organ-specific and developmental stage specific. GmHXK2 not only regulates the synthesis of AsA and the synthesis and distribution of auxin, but also promotes root elongation and induces lateral root formation, potentially enhancing soil water absorption. This study reveals the crosstalk between sugar signaling and hormone signaling in plants, where GmHXK2 acts as a glucose sensor through its interaction with GmPMM, and sheds light on the molecular mechanism by which GmHXK2 gene is involved in salt tolerance in plants.

Water deficit response in nodulated soybean roots: a comprehensive transcriptome and translatome network analysis.

BACKGROUND: Soybean establishes a mutualistic interaction with nitrogen-fixing rhizobacteria, acquiring most of its nitrogen requirements through symbiotic nitrogen fixation. This crop is susceptible to water deficit; evidence suggests that its nodulation status-whether it is nodulated or not-can influence how it responds to water deficit. The translational control step of gene expression has proven relevant in plants subjected to water deficit. RESULTS: Here, we analyzed soybean roots' differential responses to water deficit at transcriptional, translational, and mixed (transcriptional + translational) levels. Thus, the transcriptome and translatome of four combined-treated soybean roots were analyzed. We found hormone metabolism-related genes among the differentially expressed genes (DEGs) at the translatome level in nodulated and water-restricted plants. Also, weighted gene co-expression network analysis followed by differential expression analysis identified gene modules associated with nodulation and water deficit conditions. Protein-protein interaction network analysis was performed for subsets of mixed DEGs of the modules associated with the plant responses to nodulation, water deficit, or their combination. CONCLUSIONS: Our research reveals that the stand-out processes and pathways in the before-mentioned plant responses partially differ; terms related to glutathione metabolism and hormone signal transduction (2 C protein phosphatases) were associated with the response to water deficit, terms related to transmembrane transport, response to abscisic acid, pigment metabolic process were associated with the response to nodulation plus water deficit. Still, two processes were common: galactose metabolism and branched-chain amino acid catabolism. A comprehensive analysis of these processes could lead to identifying new sources of tolerance to drought in soybean.

Variability in soybean yield responses to elevated atmospheric CO2: Insights from non-structural carbohydrate remobilisation during seed filling.

The increasing atmospheric CO2 concentration (e[CO2]) has mixed effects on soybean most varieties' yield. This study elucidated the effect of e[CO2] on soybean yield and the underlying mechanisms related to photosynthetic capacity, non-structural carbohydrate (NSC) accumulation, and remobilisation. Four soybean cultivars were cultivated in open-top chambers at two CO2 levels. Photosynthesis rates were determined from R2 to R6. Plants were sampled at R5 and R8 to determine carbohydrate concentrations. There were significant variations in yield responses among the soybean cultivars under e[CO2], from no change in DS1 to a 22% increase in SN14. DS1 and SN14 had the smallest and largest increase, respectively, in daily carbon assimilation capacity. Under e[CO2], DS1, MF5, and XHJ had an increase in Ci, at which point the transition from Rubisco-limited to ribulose-1,5-bisphosphate regeneration-limited photosynthesis occurred, in contrast with SN14. Thus, the cultivars might have distinct mechanisms that enhance photosynthesis under e[CO2] conditions. A positive correlation was between daily carbon assimilation response to e[CO2] and soybean yield, emphasising the importance of enhanced photosynthate accumulation before the R5 stage in determining yield response to e[CO2]. E[CO2] significantly influenced NSC accumulation in vegetative organs at R5, with variation among cultivars. There was enhanced NSC remobilisation during seed filling, indicating cultivar-specific responses to the remobilisation of sucrose and soluble sugars, excluding sucrose and starch. A positive correlation was between leaf and stem NSC remobilisation and yield response to e[CO2], emphasising the role of genetic differences in carbohydrate remobilisation mechanisms in determining soybean yield variation under elevated CO2 levels.