Natural variation of GmFATA1B regulates seed oil content and composition in soybean.

Soybean (Glycine max) produces seeds that are rich in unsaturated fatty acids and is an important oilseed crop worldwide. Seed oil content and composition largely determine the economic value of soybean. Due to natural genetic variation, seed oil content varies substantially across soybean cultivars. Although much progress has been made in elucidating the genetic trajectory underlying fatty acid metabolism and oil biosynthesis in plants, the causal genes for many quantitative trait loci (QTLs) regulating seed oil content in soybean remain to be revealed. In this study, we identified GmFATA1B as the gene underlying a QTL that regulates seed oil content and composition, as well as seed size in soybean. Nine extra amino acids in the conserved region of GmFATA1B impair its function as a fatty acyl-acyl carrier protein thioesterase, thereby affecting seed oil content and composition. Heterogeneously overexpressing the functional GmFATA1B allele in Arabidopsis thaliana increased both the total oil content and the oleic acid and linoleic acid contents of seeds. Our findings uncover a previously unknown locus underlying variation in seed oil content in soybean and lay the foundation for improving seed oil content and composition in soybean.

Transgenic soybean of GsMYB10 shapes rhizosphere microbes to promote resistance to aluminum (Al) toxicity.

Plant resistance genes could affect rhizosphere microbiota, which in turn enhanced plant resistance to stresses. Our previous study found that overexpression of the GsMYB10 gene led to enhanced tolerance of soybean plants to aluminum (Al) toxicity. However, whether GsMYB10 gene could regulate rhizosphere microbiota to mitigate Al toxicity remains unclear. Here, we analyzed the rhizosphere microbiomes of HC6 soybean (WT) and transgenic soybean (trans-GsMYB10) at three Al concentrations, and constructed three different synthetic microbial communities (SynComs), including bacterial, fungal and cross-kingdom (bacteria and fungi) SynComs to verify their role in improving Al tolerance of soybean. Trans-GsMYB10 shaped the rhizosphere microbial communities and harbored some beneficial microbes, such as Bacillus, Aspergillus and Talaromyces under Al toxicity. Fungal and cross-kingdom SynComs showed a more effective role than the bacterial one in resistance to Al stress, and these SynComs helped soybean resist Al toxicity via affecting some functional genes that involved cell wall biosynthesis and organic acid transport etc. Overall, this study reveals the mechanism of soybean functional genes regulating the synergistic resistance of rhizosphere microbiota and plants to Al toxicity, and also highlights the possibility of focusing on the rhizobial microbial community as a potential molecular breeding target to produce crops.

GmGSTU23 Encoding a Tau Class Glutathione S-Transferase Protein Enhances the Salt Tolerance of Soybean (Glycine max L.).

Salt stress has a detrimental impact on crop yield, quality, and profitability. The tau-like glutathione transferases (GSTs) represent a significant group of enzymes that play a crucial role in plant stress responses, including salt stress. In this study, we identified a tau-like glutathione transferase family gene from soybean named GmGSTU23. Expression pattern analysis revealed that GmGSTU23 was predominantly expressed in the roots and flowers and exhibited a concentration-time-specific pattern in response to salt stress. Transgenic lines were generated and subjected to phenotypic characterization under salt stress. The transgenic lines exhibited increased salt tolerance, root length, and fresh weight compared to the wild type. Antioxidant enzyme activity and malondialdehyde content were subsequently measured, and the data revealed no significant differences between the transgenic and wild-type plants in the absence of salt stress. However, under salt stress, the wild-type plants exhibited significantly lower activities of SOD, POD, and CAT than the three transgenic lines, whereas the activity of APX and the content of MDA showed the opposite trend. We identified changes in glutathione pools and associated enzyme activity to gain insights into the underlying mechanisms of the observed phenotypic differences. Notably, under salt stress, the transgenic Arabidopsis's GST activity, GR activity, and GSH content were significantly higher than those of the wild type. In summary, our findings suggest that GmGSTU23 mediates the scavenging of reactive oxygen species and glutathione by enhancing the activity of glutathione transferase, thereby conferring enhanced tolerance to salt stress in plants.

Overexpression of GmWRKY172 enhances cadmium tolerance in plants and reduces cadmium accumulation in soybean seeds.

Introduction: Cadmium (Cd) stress is a significant threat to soybean production, and enhancing Cd tolerance in soybean is the focus of this study. The WRKY transcription factor family is associated with abiotic stress response processes. In this study, we aimed to identify a Cd-responsive WRKY transcription factor GmWRKY172 from soybean and investigate its potential for enhancing Cd tolerance in soybean. Methods: The characterization of GmWRKY172 involved analyzing its expression pattern, subcellular localization, and transcriptional activity. To assess the impact of GmWRKY172, transgenic Arabidopsis and soybean plants were generated and examined for their tolerance to Cd and Cd content in shoots. Additionally, transgenic soybean plants were evaluated for Cd translocation and various physiological stress indicators. RNA sequencing was performed to identify the potential biological pathways regulated by GmWRKY172. Results: GmWRKY172 was significantly upregulated by Cd stress, highly expressed in leaves and flowers, and localized to the nucleus with transcriptional activity. Transgenic plants overexpressing GmWRKY172 showed enhanced Cd tolerance and reduced Cd content in shoots compared to WT. Lower Cd translocation from roots to shoots and seeds was also observed in transgenic soybean. Under Cd stress, transgenic soybean accumulated less malondialdehyde (MDA) and hydrogen peroxide (H2O2) than WT plants, with higher flavonoid and lignin contents, and peroxidase (POD) activity. RNA sequencing analysis revealed that many stress-related pathways were regulated by GmWRKY172 in transgenic soybean, including flavonoid biosynthesis, cell wall synthesis, and peroxidase activity. Discussion: Our findings demonstrated that GmWRKY172 enhances Cd tolerance and reduces seed Cd accumulation in soybean by regulating multiple stress-related pathways, and could be a promising candidate for breeding Cd-tolerant and low Cd soybean varieties.

MOTHER-OF-FT-AND-TFL1 regulates the seed oil and protein content in soybean.

Soybean is a major crop that produces valuable seed oil and protein for global consumption. Seed oil and protein are regulated by complex quantitative trait loci (QTLs) and have undergone intensive selections during the domestication of soybean. It is essential to identify the major genetic components and understand their mechanism behind seed oil and protein in soybean. We report that MOTHER-OF-FT-AND-TFL1 (GmMFT) is the gene of a classical QTL that has been reported to regulate seed oil and protein content in many studies. Mutation of MFT decreased seeds oil content and weight in both Arabidopsis and soybean, whereas increased expression of GmMFT enhanced seeds oil content and weight. Haplotype analysis showed that GmMFT has undergone selection, which resulted in the extended haplotype homozygosity in the cultivated soybean and the enriching of the oil-favorable allele in modern soybean cultivars. This work unraveled the GmMFT-mediated mechanism regulating seed oil and protein content and seed weight, and revealed a previously unknown function of MFT that provides new insights into targeted soybean improvement and breeding.

GmWRKY21, a Soybean WRKY Transcription Factor Gene, Enhances the Tolerance to Aluminum Stress in Arabidopsis thaliana.

The WRKY transcription factors (TFs) are one of the largest families of TFs in plants and play multiple roles in plant growth and development and stress response. In this study, GmWRKY21 encoding a WRKY transcription factor was functionally characterized in Arabidopsis and soybean. The GmWRKY21 protein containing a highly conserved WRKY domain and a C2H2 zinc-finger structure is located in the nucleus and has the characteristics of transcriptional activation ability. The GmWRKY21 gene presented a constitutive expression pattern rich in the roots, leaves, and flowers of soybean with over 6-fold of relative expression levels and could be substantially induced by aluminum stress. As compared to the control, overexpression of GmWRKY21 in Arabidopsis increased the root growth of seedlings in transgenic lines under the AlCl3 concentrations of 25, 50, and 100 µM with higher proline and lower MDA accumulation. The results of quantitative real-time polymerase chain reaction (qRT-PCR) showed that the marker genes relative to aluminum stress including ALMT, ALS3, MATE, and STOP1 were induced in GmWRKY21 transgenic plants under AlCl3 treatment. The stress-related genes, such as KIN1, COR15A, COR15B, COR47, GLOS3, and RD29A, were also upregulated in GmWRKY21 transgenic Arabidopsis under aluminum stress. Similarly, stress-related genes, such as GmCOR47, GmDREB2A, GmMYB84, GmKIN1, GmGST1, and GmLEA, were upregulated in hair roots of GmWRKY21 transgenic plants. In summary, these results suggested that the GmWRKY21 transcription factor may promote the tolerance to aluminum stress mediated by the pathways regulating the expression of the acidic aluminum stress-responsive genes and abiotic stress-responsive genes.

GmWRKY81 Encoding a WRKY Transcription Factor Enhances Aluminum Tolerance in Soybean.

Aluminum (Al) toxicity is an essential factor that adversely limits soybean (Glycine max (L.) Merr.) growth in acid soils. WRKY transcription factors play important roles in soybean responses to abiotic stresses. Here, GmWRKY81 was screened from genes that were differentially expressed under Al treatment in Al-tolerant soybean Baxi10 and Al-sensitive soybean Bendi2. We found that GmWRKY81 was significantly induced by 20 µM AlCl3 and upregulated by AlCl3 treatment for 2 h. In different tissues, the expression of GmWRKY81 was differentially induced. In 0-1 cm root tips, the expression of GmWRKY81 was induced to the highest level. The overexpression of GmWRKY81 in soybean resulted in higher relative root elongation, root weight, depth, root length, volume, number of root tips and peroxidase activity but lower root average diameter, malonaldehyde and H2O2 contents, indicating enhanced Al tolerance. Moreover, RNA-seq identified 205 upregulated and 108 downregulated genes in GmWRKY81 transgenic lines. Fifteen of these genes that were differentially expressed in both AlCl3-treated and GmWRKY81-overexpressing soybean had the W-box element, which can bind to the upstream-conserved WRKY domain. Overall, the combined functional analysis indicates that GmWRKY81 may improve soybean Al tolerance by regulating downstream genes participating in Al3+ transport, organic acid secretion and antioxidant reactions.

Silicon-enhanced tolerance to cadmium toxicity in soybean by enhancing antioxidant defense capacity and changing cadmium distribution and transport.

Cadmium (Cd) is a widely distributed heavy metal that is toxic to plants and humans. Although silicon (Si) has been reported to reduce Cd accumulation and toxicity in plants, evidence on the functions of Si and its mechanisms in the possible alleviation of soybean are limited. Therefore, a controlled experiment was conducted to investigate the impacts and mechanisms of Si on Cd retention in soybean. Here, we determined the growth index, Cd distribution, and antioxidant activity systems of Si, as well as expression levels of differentially expressed genes (DEGs) in Si under Cd stress, and conducted RNA-seq analysis. We not only found that Si can significantly promote soybean plant growth, increase plant antioxidant activities, and reduce the Cd translocation factor, but also revealed that a total of 636 DEGs were shared between CK and Cd, CK and Cd + Si, and Cd and Cd + Si. Moreover, several genes were significantly enriched in antioxidant systems and Cd distribution and transport systems. Therefore, the expression status of Si-mediated Cd stress response genes is likely involved in improving oxidative stress and changing Cd uptake and transport, as well as improving plant growth that contributes to Si alleviating Cd toxicity in plants. Moreover, numerous potential target genes were identified for the engineering of Cd-tolerant cultivars in soybean breeding programs.

GsERF1 enhances Arabidopsis thaliana aluminum tolerance through an ethylene-mediated pathway.

Ethylene response factor (ERF) transcription factors constitute a subfamily of the AP2/ERF superfamily in plants and play multiple roles in plant growth and development as well as in stress responses. In this study, the GsERF1 gene from the wild soybean BW69 line (an Al-resistant Glycine soja line) was rapidly induced in response to aluminum stress. Quantitative real-time PCR (qRT-PCR) analysis showed that the GsERF1 gene maintained a constitutive expression pattern and was induced in soybean in response to aluminum stress, with increased amounts of transcripts detected in the roots. The putative GsERF1 protein, which contains an AP2 domain, was located in the nucleus and maintained transactivation activity. In addition, under AlCl3 treatment, GsERF1 overexpression increased the relative growth rate of the roots of Arabidopsis and weakened the hematoxylin staining of hairy roots. Ethylene synthesis-related genes such as ACS4, ACS5 and ACS6 were upregulated in GsERF1 transgenic lines compared with the wild type under AlCl3 treatment. Furthermore, the expression levels of stress/ABA-responsive marker genes, including ABI1, ABI2, ABI4, ABI5 and RD29B, in the GsERF1 transgenic lines were affected by AlCl3 treatment, unlike those in the wild type. Taken together, the results indicated that overexpression of GsERF1 may enhance aluminum tolerance of Arabidopsis through an ethylene-mediated pathway and/or ABA signaling pathway, the findings of which lay a foundation for breeding soybean plants tolerant to aluminum stress.

Identification of quantitative trait loci (QTLs) and candidate genes of seed Iron and zinc content in soybean [Glycine max (L.) Merr.].

BACKGROUND: Deciphering the hereditary mechanism of seed iron (Fe) and zinc (Zn) content in soybean is important and sustainable to address the "hidden hunger" that presently affects approximately 2 billion people worldwide. Therefore, in order to detect genomic regions related to soybean seed Fe and Zn content, a recombinant inbred line (RIL) population with 248 lines was assessed in four environments to detect Quantitative Trait Loci (QTLs) related to soybean seed Fe and Zn content. RESULT: Wide variation was found in seed Fe and Zn content in four environments, and genotype, environment, and genotype × environment interactions had significant influences on both the seed Fe and Zn content. A positive correlation was observed between seed Fe content and seed Zn content, and broad-sense heritability (H2) of seed Fe and Zn content were 0.73 and 0.75, respectively. In this study, five QTLs for seed Fe content were detected with 4.57 - 32.71% of phenotypic variation explained (PVE) and logarithm of odds (LOD) scores ranging from 3.60 to 33.79. Five QTLs controlling the seed Zn content were detected, and they individually explained 3.35 to 26.48% of the phenotypic variation, with LOD scores ranging from 3.64 to 20.4. Meanwhile, 409,541 high-quality single-nucleotide variants (SNVs) and 85,102 InDels (except intergenic regions) between two bi-parental lines were identified by whole genome resequencing. A total of 12 candidate genes were reported in one major QTL for seed Fe content and two major QTLs for seed Zn content, with the help of RNA-Seq analysis, gene ontology (GO) enrichment, gene annotation, and bi-parental whole genome sequencing (WGS) data. CONCLUSIONS: Limited studies were performed about microelement of soybean, so these results may play an important role in the biofortification of Fe and Zn and accelerate the development of marker-assisted selection (MAS) for breeding soybeans fortified with iron and zinc.

Wild Soybean Oxalyl-CoA Synthetase Degrades Oxalate and Affects the Tolerance to Cadmium and Aluminum Stresses.

Acyl activating enzyme 3 (AAE3) was identified as being involved in the acetylation pathway of oxalate degradation, which regulates the responses to biotic and abiotic stresses in various higher plants. Here, we investigated the role of Glycine sojaAAE3 (GsAAE3) in Cadmium (Cd) and Aluminum (Al) tolerances. The recombinant GsAAE3 protein showed high activity toward oxalate, with a Km of 105.10 ± 12.30 µM and Vmax of 12.64 ± 0.34 µmol min-1 mg-1 protein, suggesting that it functions as an oxalyl-CoA synthetase. The expression of a GsAAE3-green fluorescent protein (GFP) fusion protein in tobacco leaves did not reveal a specific subcellular localization pattern of GsAAE3. An analysis of the GsAAE3 expression pattern revealed an increase in GsAAE3 expression in response to Cd and Al stresses, and it is mainly expressed in root tips. Furthermore, oxalate accumulation induced by Cd and Al contributes to the inhibition of root growth in wild soybean. Importantly, GsAAE3 overexpression increases Cd and Al tolerances in A. thaliana and soybean hairy roots, which is associated with a decrease in oxalate accumulation. Taken together, our data provide evidence that the GsAAE3-encoded protein plays an important role in coping with Cd and Al stresses.

High Aluminum Drives Different Rhizobacterial Communities Between Aluminum-Tolerant and Aluminum-Sensitive Wild Soybean.

Aluminum (Al)-resistant plant cultivars can recruit beneficial microbes to alleviate the stresses. However, the mechanism of how rhizobacterial communities strengthen Al tolerance of wild soybean has not been addressed. The aim of this study was to investigate the bacterial community structure in the rhizosphere of Al-tolerant (BW69) and Al-sensitive (W270) wild soybean germplasm subjected to three Al concentrations. We analyzed the rhizobacterial communities of the two genotypes by high-throughput sequencing of 16S rRNA genes. The results showed that high Al stress recruited different rhizobacterial communities between two genotypes. In total, 49 OTUs, such as OTU15 (Gammaproteobacteria_KF-JG30-C25_norank), OTU23 (Mizugakiibacter), and OTU93 (Alkanibacter), were enriched in the rhizosphere of BW69 at the low and high Al concentrations. Moreover, bacterial community in the rhizosphere of BW69 had a more complex co-occurrence network than did W270 at the high Al concentration. Overall, our findings highlighted that high Al concentration magnified the difference in rhizobacterial community structure between two genotypes. However, the lower modularity of the co-occurrence network in rhizosphere of BW69 than W270 under Al stress may cause the rhizobacterial community to be less resistant and more influenced by disturbance. This study emphasizes the possibility of using rhizobacteria as an improved crop breeding or gene to produce crops that are more resistant to the toxicity of heavy metal.

Rhizosphere Soil Fungal Communities of Aluminum-Tolerant and -Sensitive Soybean Genotypes Respond Differently to Aluminum Stress in an Acid Soil.

Different soybean genotypes can differ in their tolerance toward aluminum stress depending on their rhizosphere-inhabiting microorganisms. However, there is limited understanding of the response of fungal communities to different aluminum concentrations across different genotypes. Here, we used metabarcoding of fungal ribosomal markers to assess the effects of aluminum stress on the rhizosphere fungal community of aluminum-tolerant and aluminum-sensitive soybean genotypes. Shifts in fungal community structure were related to changes in plant biomass, fungal abundance and soil chemical properties. Aluminum stress increased the difference in fungal community structure between tolerant and sensitive genotypes. Penicillium, Cladosporium and Talaromyces increased with increasing aluminum concentration. These taxa associated with the aluminum-tolerant genotypes were enriched at the highest aluminum concentration. Moreover, complexity of the co-occurrence network associated with the tolerant genotypes increased at the highest aluminum concentration. Collectively, increasing aluminum concentrations magnified the differences in fungal community structure between the two studied tolerant and sensitive soybean genotypes. This study highlights the possibility to focus on rhizosphere fungal communities as potential breeding target to produce crops that are more tolerant toward heavy metal stress or toxicity in general.

Transcription Factor GmWRKY142 Confers Cadmium Resistance by Up-Regulating the Cadmium Tolerance 1-Like Genes.

Cadmium (Cd) is a widespread pollutant that is toxic to living organisms. Previous studies have identified certain WRKY transcription factors, which confer Cd tolerance in different plant species. In the present study, we have identified 29 Cd-responsive WRKY genes in Soybean [Glycine max (L.) Merr.], and confirmed that 26 of those GmWRKY genes were up-regulated, while 3 were down-regulated. We have also cloned the novel, positively regulated GmWRKY142 gene from soybean and investigated its regulatory mechanism in Cd tolerance. GmWRKY142 was highly expressed in the root, drastically up-regulated by Cd, localized in the nucleus, and displayed transcriptional activity. The overexpression of GmWRKY142 in Arabidopsis thaliana and soybean hairy roots significantly enhanced Cd tolerance and lead to extensive transcriptional reprogramming of stress-responsive genes. ATCDT1, GmCDT1-1, and GmCDT1-2 encoding cadmium tolerance 1 were induced in overexpression lines. Further analysis showed that GmWRKY142 activated the transcription of ATCDT1, GmCDT1-1, and GmCDT1-2 by directly binding to the W-box element in their promoters. In addition, the functions of GmCDT1-1 and GmCDT1-2, responsible for decreasing Cd uptake, were validated by heterologous expression in A. thaliana. Our combined results have determined GmWRKYs to be newly discovered participants in response to Cd stress, and have confirmed that GmWRKY142 directly targets ATCDT1, GmCDT1-1, and GmCDT1-2 to decrease Cd uptake and positively regulate Cd tolerance. The GmWRKY142-GmCDT1-1/2 cascade module provides a potential strategy to lower Cd accumulation in soybean.

Identification and Mapping of Stable QTLs for Seed Oil and Protein Content in Soybean [Glycine max (L.) Merr.].

This research aimed to identify stable quantitative trait loci (QTL) associated with oil and protein content in soybean. A population of 196 recombinant inbred lines (RILs) derived from Huachun 2 × Wayao was used to evaluate these target traits. A high-density genetic linkage map was constructed by using high-throughput genome-wide sequencing technology, which contained 3413 recombination bin markers and spanned 5400.4 cM with an average distance of 1.58 cM between markers. Eighteen stable QTLs controlling oil and protein content were detected. Among them, qOil-11-1 was identified for the first time as a novel QTL, while qOil-5-1, qPro-10-1, and qPro-14-1 were strong and stable QTLs with high log-likelihood (LOD) values. Sixteen differentially expressed genes (DEGs) within these four QTLs were shown to be highly expressed during seed development based on RNA sequencing (RNA-seq) data analysis. Our results may contribute toward gene mining and marker-assisted selection (MAS) for breeding a high-quality soybean in the future.

GsMAS1 Encoding a MADS-box Transcription Factor Enhances the Tolerance to Aluminum Stress in Arabidopsis thaliana.

The MADS-box transcription factors (TFs) are essential in regulating plant growth and development, and conferring abiotic and metal stress resistance. This study aims to investigate GsMAS1 function in conferring tolerance to aluminum stress in Arabidopsis. The GsMAS1 from the wild soybean BW69 line encodes a MADS-box transcription factor in Glycine soja by bioinformatics analysis. The putative GsMAS1 protein was localized in the nucleus. The GsMAS1 gene was rich in soybean roots presenting a constitutive expression pattern and induced by aluminum stress with a concentration-time specific pattern. The analysis of phenotypic observation demonstrated that overexpression of GsMAS1 enhanced the tolerance of Arabidopsis plants to aluminum (Al) stress with larger values of relative root length and higher proline accumulation compared to those of wild type at the AlCl3 treatments. The genes and/or pathways regulated by GsMAS1 were further investigated under Al stress by qRT-PCR. The results indicated that six genes resistant to Al stress were upregulated, whereas AtALMT1 and STOP2 were significantly activated by Al stress and GsMAS1 overexpression. After treatment of 50 µM AlCl3, the RNA abundance of AtALMT1 and STOP2 went up to 17-fold and 37-fold than those in wild type, respectively. Whereas the RNA transcripts of AtALMT1 and STOP2 were much higher than those in wild type with over 82% and 67% of relative expression in GsMAS1 transgenic plants, respectively. In short, the results suggest that GsMAS1 may increase resistance to Al toxicity through certain pathways related to Al stress in Arabidopsis.

Characterization of the Soybean GmIREG Family Genes and the Function of GmIREG3 in Conferring Tolerance to Aluminum Stress.

The IREG (IRON REGULATED/ferroportin) family of genes plays vital roles in regulating the homeostasis of iron and conferring metal stress. This study aims to identify soybean IREG family genes and characterize the function of GmIREG3 in conferring tolerance to aluminum stress. Bioinformatics and expression analyses were conducted to identify six soybean IREG family genes. One GmIREG, whose expression was significantly enhanced by aluminum stress, GmIREG3, was studied in more detail to determine its possible role in conferring tolerance to such stress. In total, six potential IREG-encoding genes with the domain of Ferroportin1 (PF06963) were characterized in the soybean genome. Analysis of the GmIREG3 root tissue expression patterns, subcellular localizations, and root relative elongation and aluminum content of transgenic Arabidopsis overexpressing GmIREG3 demonstrated that GmIREG3 is a tonoplast localization protein that increases transgenic Arabidopsis aluminum resistance but does not alter tolerance to Co and Ni. The systematic analysis of the GmIREG gene family reported herein provides valuable information for further studies on the biological roles of GmIREGs in conferring tolerance to metal stress. GmIREG3 contributes to aluminum resistance and plays a role similar to that of FeIREG3. The functions of other GmIREG genes need to be further clarified in terms of whether they confer tolerance to metal stress or other biological functions.

GsMATE encoding a multidrug and toxic compound extrusion transporter enhances aluminum tolerance in Arabidopsis thaliana.

BACKGROUND: Multidrug and toxic compound extrusion (MATE) transporters, which exist widely in plants, function as crucial regulators in plant resistance to aluminum (Al) toxicity by inducing citrate efflux. However, the functions of most MATE family members in soybean (Glycine soja) remain to be elucidated. RESULTS: Expression pattern analysis showed that GsMATE was constitutively expressed in different soybean organs, with the highest level in root compared with those in stem, leaf and cotyledon. In addition, Al stress induced expression of GsMATE in soybean. Temporal analysis indicated that GsMATE expression was greatly enhanced by increasing concentrations of aluminum [Al3+] after short exposure, reaching the high levels detected in the BW69 (Al-resistant) and the JW81 (Al-sensitive) lines of Glycine soja of wild soybean at 6 h and 8 h, respectively. Furthermore, transient GsMATE expression in Arabidopsis protoplasts showed that GsMATE protein localized to the plasma membrane. Overexpression of GsMATE on an Arabidopsis columbia-0 (Col-0) background resulted in increased Al tolerance in transgenic plants. Analysis of hematoxylin staining showed that the roots of GsMATE transgenic lines were stained less intensely than those of the wild-type exposured to the same AlCl3 concentrations. Therefore, GsMATE enhanced the resistance of transgenic plants to Al toxicity by reducing Al accumulation in Arabidopsis roots. CONCLUSIONS: In summary, our results indicate that GsMATE is responsive to aluminum stress and may participate in the regulation of sensitivity to Al toxicity in Arabidopsis. In addition, the GsMATE protein is an Al-induced citrate transporter of the MATE family and exerts an essential role in Al tolerance in Glycine soja.