Tartary buckwheat rutin: Accumulation, metabolic pathways, regulation mechanisms, and biofortification strategies.

Rutin is a significant flavonoid with strong antioxidant property and various therapeutic effects. It plays a crucial role in disease prevention and human health maintenance, especially in anti-inflammatory, antidiabetic, hepatoprotective and cardiovascular effects. While many plants can synthesize and accumulate rutin, tartary buckwheat is the only food crop possessing high levels of rutin. At present, the rutin content (RC) is regarded as the key index for evaluating the nutritional quality of tartary buckwheat. Consequently, rutin has become the focus for tartary buckwheat breeders and has made considerable progress. Here, we summarize research on the rutin in tartary buckwheat in the past two decades, including its accumulation, biosynthesis and breakdown pathways, and regulatory mechanisms. Furthermore, we propose several strategies to increase the RC in tartary buckwheat seeds based on current knowledge. This review aims to provide valuable references for elevating the quality of tartary buckwheat in the future.

Accumulation of the bitter substance quercetin mediated by the overexpression of a novel seed-specific gene FtRDE2 in Tartary buckwheat.

Tartary buckwheat (Fagopyrum tataricum) is frequently employed as a resource to develop health foods, owing to its abundant flavonoids such as rutin. However, the consumption of Tartary buckwheat (TB) is limited in food products due to the strong bitterness induced by the hydrolysis of rutin into quercetin. This transformation is facilitated by the degrading enzyme (RDE). While multiple RDE isoenzymes exist in TB, the superior coding gene of FtRDEs has not been fully explored, which hinders the breeding of TB varieties with minimal bitterness. Here, we found that FtRDE2 is the most abundant enzyme in RDE crude extracts, and its corresponding gene is specifically expressed in TB seeds. Results showed that FtRDE2 has strong rutin hydrolysis activity. Overexpression of FtRDE2 not only significantly promoted rutin hydrolysis and quercetin accumulation but also dramatically upregulated genes involved in the early phase of flavonoid synthesis (FtPAL1、FtC4H1、Ft4CL1, FtCHI1) and anthocyanin metabolism (FtDFR1). These findings elucidate the role of FtRDE2, emphasizing it as an endogenous factor contributing to the bitterness in TB and its involvement in the metabolic regulatory network. Moreover, correlation analysis revealed a positive relationship between the catalytic activity of RDE extracts and the expression level of FtRDE2 during seed germination. In summary, our results suggest that FtRDE2 can serve as a promising candidate for the molecular breeding of a TB variety with minimal bitterness.

The Tartary buckwheat bHLH gene ALCATRAZ contributes to silique dehiscence in Arabidopsis thaliana.

Tartary buckwheat is popular because of its rich nutrients. However, the difficulty in shelling restricts food production. The gene ALCATRAZ (AtALC) plays a key role in silique dehiscence in Arabidopsis thaliana. In this study, an atalc mutant was obtained by CRISPR/Cas9, and a FtALC gene homologous to AtALC was complemented into the atalc mutant to verify its function. Phenotypic observations showed that three atalc mutant lines did not dehiscence, while ComFtALC lines recovered the dehiscence phenotype. The contents of lignin, cellulose, hemicellulose, and pectin in the siliques of all the atalc mutant lines were significantly higher than those in the wild-type and ComFtALC lines. Moreover, FtALC was found to regulate the expression of cell wall pathway genes. Finally, the interaction of FtALC with FtSHP and FtIND was verified by yeast two-hybrid, bimolecular fluorescent complimentary (BIFC) and firefly luciferase completion imaging assays (LCIs). Our findings enrich the silique regulatory network and lay the foundation for the cultivation of easily shelled tartary buckwheat varieties.

Genome-Wide Identification of ATG Gene Family Members in Fagopyrum tataricum and Their Expression during Stress Responses

Abiotic stresses such as drought and salinity are major environmental factors limiting plant productivity. Autophagy-related genes are extensively involved in plant growth, development, and adverse stress responses, which have not yet been characterized in Tartary buckwheat (Fagopyrum tataricum, TB). In this study, we verified that drought stress could induce autophagy in TB roots. Next, 49 FtATGs in the whole genome of TB were identified. All FtATGs were randomly distributed in 8 known chromosomes, while 11 FtATGs were predictably segmental repeats. As the core component of autophagy, there were 8 FtATG8s with similar gene structures in TB, while FtATG8s showed high expression at the transcription level under drought and salt stresses. The cis-acting element analysis identified that all FtATG8 promoters contain light-responsive and MYB-binding elements. FtATG8s showed a cell-wide protein interaction network and strongly correlated with distinct stress-associated transcription factors. Furthermore, overexpression of FtATG8a and FtATG8f enhanced the antioxidant enzyme activities of TB under adverse stresses. Remarkably, FtATG8a and FtATG8f may be vital candidates functioning in stress resistance in TB. This study prominently aids in understanding the biological role of FtATG genes in TB.

A Novel R2R3-MYB Transcription Factor FtMYB22 Negatively Regulates Salt and Drought Stress through ABA-Dependent Pathway.

Tartary buckwheat (Fagopyrum tataricum Gaertn.) is a coarse cereal with strongly abiotic resistance. The MYB family plays a regulatory role in plant growth, development, and responses to biotic and abiotic stresses. However, the characteristics and regulatory mechanisms of MYB transcription factors in Tartary buckwheat remain unclarified. Here, this study cloned the FtMYB22 gene from Tartary buckwheat, and investigated its involvement in responding to individual water deficit and salt stress in Arabidopsis. Sequence analysis highlighted that the N-termini of FtMYB22 contained two highly conserved SANT domains and one conserved domain from the SG20 subfamily. Nucleus-localized FtMYB22 did not have individual transcriptional activation activity. Water deficiency and salt stress induced the high expression of the GUS gene, which was driven by the promoter of FtMYB22. Yeast stress experiments showed that the overexpression of FtMYB22 significantly reduced the growth activity of transgenic yeast under water deficit or salt stress. Consistently, the overexpression of FtMYB22 reduced the salt and water deficit stress resistance of the transgenic plants. In addition, physiological parameters showed that transgenic plants had lower proline and antioxidant enzyme activity under stress conditions. Compared to the wild-type (WT), transgenic plants accumulated more malondialdehyde (MDA), H2O2, and O2−; they also showed higher ion permeability and water loss rates of detached leaves under stress treatments. Notably, FtMYB22 was involved in plant stress resistance through an ABA-dependent pathway. Under stress conditions, the expression of RD29A, RD29B, PP2CA, KIN1, COR15A, and other genes in response to plant stress in transgenic lines was significantly lower than that in the WT (p < 0.05). Furthermore, yeast two-hybrid assay showed that there was a significant interaction between FtMYB22 and the ABA receptor protein RCAR1/2, which functioned in the ABA signal pathway. Altogether, FtMYB22, as a negative regulator, inhibited a variety of physiological and biochemical reactions, affected gene expression and stomatal closure in transgenic plants through the ABA-dependent pathway, and reduced the tolerance of transgenic Arabidopsis to water deficiency and salt stress. Based on these fundamental verifications, further studies would shed light on the hormone signal response mechanism of FtMYB22.

Transcriptomic and Metabonomic Profiling Reveals the Antihyperlipidemic Effects of Tartary Buckwheat Sprouts in High-Fat-Diet-Fed Mice.

Flavonoids are known for potent antioxidant activity and antihyperlipidemia. As a result of the few antinutritional factors and high bioactive substances, such as flavonoids, sprouts of tartary buckwheat (Fagopyrum tataricum, STB) have become healthy food. This study aims to unravel the antihyperlipidemic effects of STB in vivo and its potential mechanism through transcriptomic and metabonomic analysis. The physiological parameters of mice administered the high-fat diet with or without 2.5 and 5% of STB for 10 weeks were recorded. Liquid chromatography-tandem mass spectrometry and RNA sequencing were applied to obtain the serum lipid metabolomic and hepatic transcriptomic profiling, respectively. Results revealed that STB could significantly alleviate the increase of body weight, liver, and abdominal adipose while ameliorating the lipid content in serum and insulin resistance of mice fed with a high-fat diet. Notably, the metabonomic analysis identified the core differential metabolites mainly enriched in the pathways, such as fat digestion and absorption, insulin resistance, and other processes. Transcriptomic results revealed that STB significantly altered the expression levels of PIK3R1, LRP5, SLC10A2, and FBXO21. These genes are involved in the PI3K-AKT signaling pathway, digestion and absorption of carbohydrates, and type II diabetes mellitus pathways. In this study, STB exhibited remarkable influence on the metabolism of lipids and glucose, exerting antihyperlipidemic effects. STB have the potential for the development and application of a lipid-lowering health food.

Tartary buckwheat FtMYB30 transcription factor improves the salt/drought tolerance of transgenic Arabidopsis in an ABA-dependent manner.

Drought and high salinity affect plant growth, development, yield, and quality. MYB transcription factors (TFs) in plants play an indispensable regulatory role in resisting adverse stress. In this study, screening and functional validation of the TF FtMYB30, which can respond extensively to abiotic stress and abscisic acid (ABA), was achieved in Tartary buckwheat. FtMYB30, one of the SG22 (sub-group 22) family of R2R3-MYB TFs, localized in the nucleus and had transcriptional activation activity. Under drought and salt stress, FtMYB30 overexpression reduced the oxidative damage in transgenic plants by increasing the activity of proline content and antioxidant enzymes and significantly upregulate the expression of RD29A, RD29B, and Cu/ZnSOD, thereby enhancing drought/salt tolerance in transgenic Arabidopsis. Additionally, overexpression of FtMYB30 can reduce the sensitivity of transgenic plants to ABA. Moreover, AtRCAR1/2/3 and AtMPK6 directly interact with the FtMYB30 TF, possibly through the crosstalk between MAPKs (mitogen-activated protein kinases) and the ABA signaling pathway. Taken together, these results suggest that FtMYB30, as a positive regulator, mediates plant tolerance to salt and drought through an ABA-dependent signaling pathway.

FtNAC31, a Tartary buckwheat NAC transcription factor, enhances salt and drought tolerance in transgenic Arabidopsis.

Tartary buckwheat [Fagopyrum tataricum (L.) Gaertn.] is a pseudocereal with strongly abiotic resistance. NACs, one of the largest plant-specific transcription factors (TFs), are involved in various stress responses. However, the characteristics and regulatory mechanisms of NAC TFs remain unclarified clearly in Tartary buckwheat (TB). In this study, it validated that salt, drought, and abscisic acid (ABA) stress significantly up-regulated the expression of NAC TF gene FtNAC31. Its coding protein has a C-terminal transactivated domain and localized in the nucleus, suggesting that FtNAC31 might play a transcriptional activation role in TB. Notably, overexpression of FtNAC31 lowered the seed germination rate upon ABA treatment and enhanced the tolerance to salt and drought stress in transgenetic Arabidopsis. Furthermore, under various stresses, the activities of superoxide dismutase (SOD), peroxidase (POD), and catalase (CAT) in FtNAC31 overexpressed lines exhibited a sharp increase trend. Meanwhile, the expression levels of several stress-associated genes including RD29A, RD29B, RD22, DREB2B, NCED3, and POD1, were dramatically upregulated in lines overexpressing FtNAC31. Altogether, overproduction of FtNAC31 could enhance the resistance to salt and drought stresses in transgenic Arabidopsis, which most likely functioned in an ABA-dependent way.

Tartary Buckwheat R2R3-MYB Gene FtMYB3 Negatively Regulates Anthocyanin and Proanthocyanin Biosynthesis.

Anthocyanins and proanthocyanidins (PAs) are vital secondary metabolites in Tartary buckwheat because of their antioxidant capacities and radical scavenging functions. It has been demonstrated that R2R3-MYB transcription factors (TFs) are essential regulators of anthocyanin and PA biosynthesis in many plants. However, their regulatory mechanisms in Tartary buckwheat remain to be clarified. Here, we confirmed the role of FtMYB3 in anthocyanin and PA biosynthesis. FtMYB3, which belongs to the subgroup 4 R2R3 family was predominantly expressed in roots. The transcriptional expression of FtMYB3 increased significantly under hormone treatment with SA and MeJA and abiotic stresses including drought, salt, and cold at the seedling stage. Functional analyses showed that FtMYB3 negatively regulated anthocyanin and PA biosynthesis, primarily via downregulating the expression of the DFR, ANS, BAN, and TT13 in transgenic Arabidopsis thaliana, which may depend on the interaction between FtMYB3 and FtbHLH/FtWD40. Altogether, this study reveals that FtMYB3 is a negative regulatory transcription factor for anthocyanin and PA biosynthesis in Tartary buckwheat.

Genome-wide identification of MAPK gene family members in Fagopyrum tataricum and their expression during development and stress responses.

BACKGROUND: Mitogen-activated protein kinases (MAPKs) plays essential roles in the development, hormone regulation and abiotic stress response of plants. Nevertheless, a comprehensive study on MAPK family members has thus far not been performed in Tartary buckwheat. RESULTS: Here, we identified 16 FtMAPKs in the Fagopyrum tataricum genome. Phylogenetic analysis showed that the FtMAPK family members could be classified into Groups A, B, C and D, in which A, B and C members contain a Thr-Glu-Tyr (TEY) signature motif and Group D members contain a Thr-Asp-Tyr (TDY) signature motif. Promoter cis-acting elements showed that most ProFtMAPks contain light response elements, hormone response elements and abiotic stress response elements, and several ProFtMAPks have MYB-binding sites, which may be involved in the regulation of flavonoid biosynthesis-related enzyme gene expression. Synteny analysis indicated that FtMAPKs have a variety of biological functions. Protein interaction prediction suggested that MAPKs can interact with proteins involved in development and stress resistance. Correlation analysis further confirmed that most of the FtMAPK genes and transcription factors involved in the stress response have the same expression pattern. The transient transformation of FtMAPK1 significantly increased the antioxidant enzymes activity in Tartary buckwheat leaves. In addition, we also found that FtMAPK1 can respond to salt stress by up-regulating the transcription abundance of downstream genes. CONCLUSIONS: A total of 16 MAPKs were identified in Tartary buckwheat, and the members of the MAPK family containing the TDY motif were found to have expanded. The same subfamily members have relatively conserved gene structures and similar protein motifs. Tissue-specific expression indicated that the expression of all FtMAPK genes varied widely in the roots, stems, leaves and flowers. Most FtMAPKs can regulate the expression of other transcription factors and participate in the abiotic stress response. Our findings comprehensively revealed the FtMAPK gene family and laid a theoretical foundation for the functional characterization of FtMAPKs.

Genome-Wide Investigation of Major Enzyme-Encoding Genes in the Flavonoid Metabolic Pathway in Tartary Buckwheat (Fagopyrum tataricum).

Key enzymes play a vital role in plant growth and development. However, the evolutionary relationships between genes encoding key enzymes in the metabolic pathway of Tartary buckwheat flavonoids are poorly understood. Based on the published Tartary buckwheat genome sequence and related Tartary buckwheat transcriptome data, 48 key enzyme-encoding genes involved in flavonoid metabolism were screened from the Tartary buckwheat genome in this study; the chromosome localization, gene structure and promoter elements of these enzyme-encoding gene were also investigated. Gene structure analysis revealed relatively conserved 5' exon sequences among the 48 genes, indicating that the structural diversity of key enzyme-encoding genes is low in Tartary buckwheat. Through promoter analysis, these key enzyme-encoding genes were found to contain a large number of light-response elements and hormone-response elements. In addition, some genes could bind MYB transcription factors, participating in the regulation of flavonoid biosynthesis. The transcription level of the 48 key enzyme-encoding gene varied greatly among tissues. In this study, we identified 48 key enzyme-encoding genes involved in flavonoid metabolic pathways, and elucidated the structure, evolution and tissue-specific expression patterns of these genes. These results lay a foundation for further understanding the functional characteristics and evolutionary relationships of key enzyme-encoding genes involved in the flavonoid metabolic pathway in Tartary buckwheat.

Enhanced antioxidant capacity and upregulated transporter genes contribute to the UV-B-induced increase in blinin in Conyza blinii.

Conyza blinii (C. blinii) is a traditional Chinese medicinal plant mainly grown in Sichuan, China. C. blinii is suitable for studying the mechanism of plant tolerance to UV-B due to its living conditions, characterized by a high altitude and exposure to strong ultraviolet radiation. Our results showed that the growth and photosynthetic activity of C. blinii were improved under a specific intensity of UV-B, rather than being significantly inhibited. Although UV-B increased the content of reactive oxygen species (ROS) in C. blinii, the activities of antioxidative enzymes were elevated, including superoxide dismutase (SOD), peroxidase (POD), catalase (CAT), and ascorbate peroxidase (APX), which contributed to the elimination of ROS. Additionally, the content of blinin, the characteristic diterpene in C. blinii, was markedly increased by UV-B. Furthermore, RNA sequencing analyses were used to explore the molecular mechanism of UV-B tolerance in C. blinii. According to the results, most of the key enzyme genes in the blinin synthesis pathway were upregulated by UV-B. In addition, 23 upregulated terpene transporter genes were identified, and these genes might participate in blinin transport during the response to UV-B. Taken together, these results implied that enhanced antioxidant capacity and upregulated transporter genes contributed to increased synthesis of blinin in response to UV-B in C. blinii.

FtMYB6, a Light-Induced SG7 R2R3-MYB Transcription Factor, Promotes Flavonol Biosynthesis in Tartary Buckwheat (Fagopyrum tataricum).

Tartary buckwheat (Fagopyrum tataricum) is rich in flavonols, which are thought to be highly beneficial for human health. However, little is known about the regulatory mechanism of flavonol biosynthesis in Tartary buckwheat. In this study, we identified and characterized a novel SG7 R2R3-MYB transcription factor in Tartary buckwheat, FtMYB6. We showed that FtMYB6 is located in the nucleus and acts as a transcriptional activator. The FtMYB6 promoter showed strong spatiotemporal specificity and was induced by light. The expression of FtMYB6 showed a significant correlation with rutin accumulation in the roots, stems, leaves, and flowers. Overexpression of FtMYB6 in transgenic Tartary buckwheat hairy roots and tobacco (Nicotiana tabacum) plants significantly increased the accumulation of flavonols. In transient luciferase (LUC) activity assay, FtMYB6 promoted the activity of FtF3H and FtFLS1 promoters and inhibited the activity of the Ft4CL promoter. Collectively, our results suggest that FtMYB6 promotes flavonol biosynthesis by activating FtF3H and FtFLS1 expression.

Ferrous iron-induced increases in capitate glandular trichome density and upregulation of CbHO-1 contributes to increases in blinin content in Conyza blinii.

MAIN CONCLUSION: Ferrous iron can promote the development of glandular trichomes and increase the content of blinin, which depends on CbHO-1 expression. Conyza blinii (C. blinii) is a unique Chinese herbal medicine that grows in Sichuan Province, China. Because the habitat of C. blinii is an iron ore mining area with abundant iron content, this species can be used as one of the best materials to study the mechanism of plant tolerance to iron. In this study, C. blinii was treated with ferrous-EDTA solutions at different concentrations, and it was found that the tolerance value of C. blinii to iron was 200 µM. Under this concentration, the plant height, root length, biomass, and iron content of C. blinii increased to the maximum values, and the effect was dependent on the upregulated expression of CbHO-1. At the same time, under ferrous iron, the photosynthetic capacity and capitate glandular trichome density of C. blinii also significantly increased, providing precursors and sites for the synthesis of blinin, thus significantly increasing the content of blinin. These processes were also dependent on the high expression of CbHO-1. Correlation analysis showed that there were strong positive correlations between iron content, capitate glandular trichome density, CbHO-1 gene expression, and blinin content. This study explored the effects of ferrous iron on the physiology and biochemistry of C. blinii, greatly improving our understanding of the mechanism of iron tolerance in C. blinii.

FtMYB18 acts as a negative regulator of anthocyanin/proanthocyanidin biosynthesis in Tartary buckwheat.

KEY MESSAGE: FtMYB18 plays a role in the repression of anthocyanins and proanthocyanidins accumulation by strongly down-regulating the CHS and DFR genes in Tartary buckwheat, and the C5 motif plays an important role in this process. Anthocyanins and proanthocyanidins (PAs) are important flavonoids in Tartary buckwheat (Fagopyrum tataricum Gaertn.), which provides various vibrant color and stronge abiotic stress resistance. Their synthesis is generally regulated by MYB transcription factors at transcription level. However, the negative regulations of MYB and their effects on flavonol metabolism are poorly understood. A SG4-like MYB subfamily TF, FtMYB18, containing C5 motif was identified from Tartary buckwheat. The expression of FtMYB18 was not only showed a negative correlation with anthocyanins and PAs content but also strongly respond to MeJA and ABA. As far as the transgenic lines with FtMYB18 overexpression, anthocyanins and PAs accumulations were decreased through down-regulating expression levels of NtCHS and NtDFR in tobacco, AtDFR and AtTT12 in Arabidopsis, FtCHS, FtDFR and FtANS in Tartary buckwheat hairy roots, respectively. However, FtMYB18 showed no effect on the FLS gene expression and the metabolites content in flavonol synthesis branch. The further molecular interaction analysis indicated FtMYB18 could mediate the inhibition of anthocyanins and PAs synthesis by forming MBW transcriptional complex with FtTT8 and FtTTG1, or MYB-JAZ complex with FtJAZ1/-3/-4/-7. Importantly, in FtMYB18 mutant lines with C5 motif deletion (FtMYB18-C), both of anthocyanins and PAs accumulations had recovered to the similar level as that in wild type, which was attributed to the weakened MBW complex activity or the deficient molecular interaction between FtMYB18ΔC5 with FtJAZ3/-4. The results showed that FtMYB18 could suppress anthocyanins and PAs synthesis at transcription level through the specific interaction of C5 motif with other proteins in Tartary buckwheat.

Isolation and identification of flavonoid-producing endophytic fungi from medicinal plant Conyza blinii H.Lév that exhibit higher antioxidant and antibacterial activities.

BACKGROUND: Conyza blinii H. Lév is a medicinal plant that has a variety of pharmacological activities, but its study is at a standstill due to the shortage of resources. METHOD: This study utilized the surface sterilization method to isolate endophytic fungi, and they were preliminarily identified by morphology. Flavonoid-producing strains were screened by NaNO2-Al(NO)3 colorimetry and further identified by the ITS sequence. Additionally, we used five antioxidant assays (DPPH, Hydroxyl radical, ABTS, FRAP and T-AOC assays) to systematically evaluate the antioxidant capacity of total flavonoids , and we also determined their antibacterial activity. RESULTS: In this study, 21 endophytic fungi were isolated from wild Conyza blinii H. Lév for the first time. There were six flavonoid-producing strains, especially CBL11, whose total flavonoid content reached 50.78 ± 2.4 mg/L. CBL12, CBL12-2 and CBL1-1 all exhibited excellent antioxidant activity. The effect of CBL12 was similar to that of ascorbic acid at low concentrations, and its radical scavenging rates for DPPH and ABTS were 94.56 ± 0.29 % and 99.88 ± 0.27%, respectively, while its IC50 values were only 0.11 ± 0.01 mg/mL and 0.2 ± 0.01 mg/mL. Through LC-MS, we found that CBL12 could produce many high-value flavonoids, such as 3-methoxyflavone, nobiletin, formononetin, scopoletin, and daidzein. Additionally, CBL9 had good antibacterial activity against both gram-positive and gram-negative bacteria. Notably, we obtained the high-yield strains CBL12 and CBL9, which not only had high yields (10.64 ± 1.01 mg/L and 10.17 ± 0.11 mg/L, respectively) but also had excellent biological activity. Hence, the results of this study provide new ideas for endophytic fungi that can be exploited as a source of flavonoids and other medicinal components from Conyza blinii H. Lév. Moreover, this study can serve as a reference for the development of rare medicinal materials.

Tartary Buckwheat Transcription Factor FtbZIP5, Regulated by FtSnRK2.6, Can Improve Salt/Drought Resistance in Transgenic Arabidopsis.

bZIP transcription factors have been reported to be involved in many different biological processes in plants. The ABA (abscisic acid)-dependent AREB/ABF-SnRK2 pathway has been shown to play a key role in the response to osmotic stress in model plants. In this study, a novel bZIP gene, FtbZIP5, was isolated from tartary buckwheat, and its role in the response to drought and salt stress was characterized by transgenic Arabidopsis. We found that FtbZIP5 has transcriptional activation activity, which is located in the nucleus and specifically binds to ABRE elements. It can be induced by exposure to PEG6000, salt and ABA in tartary buckwheat. The ectopic expression of FtbZIP5 reduced the sensitivity of transgenic plants to drought and high salt levels and reduced the oxidative damage in plants by regulating the antioxidant system at a physiological level. In addition, we found that, under drought and salt stress, the expression levels of several ABA-dependent stress response genes (RD29A, RD29B, RAB18, RD26, RD20 and COR15) in the transgenic plants increased significantly compared with their expression levels in the wild type plants. Ectopic expression of FtbZIP5 in Arabidopsis can partially complement the function of the ABA-insensitive mutant abi5-1 (abscisic acid-insensitive 5-1). Moreover, we screened FtSnRK2.6, which might phosphorylate FtbZIP5, in a yeast two-hybrid experiment. Taken together, these results suggest that FtbZIP5, as a positive regulator, mediates plant tolerance to salt and drought through ABA-dependent signaling pathways.

A WRKY transcription factor, FtWRKY46, from Tartary buckwheat improves salt tolerance in transgenic Arabidopsis thaliana.

The WRKY transcription factor family includes plant-specific transcription factors that are widely involved in plant biotic and abiotic stress responses, growth and development. Tartary buckwheat is a type of small grain with strong resistance to adverse growing conditions. No systematic exploration of the WRKY family in Tartary buckwheat has yet been reported. In this paper, we report the FtWRKY46 gene from Tartary buckwheat and study its role in salt tolerance. FtWRKY46 has transcriptional activation activity in yeast, and FtWRKY46 fused to yellow fluorescent protein localizes to the nucleus. Further studies have found that its transcriptional activation region is located at the N-terminus. A yeast one-hybrid assay indicated that FtWRKY46 could bind to a W-box and activate reporter gene expression. Similarly, transient cotransfection showed that FtWRKY46 could specifically bind to W-box regions and activate reporter gene expression in plants. Furthermore, ectopic expression of FtWRKY46 could enhance Arabidopsis tolerance to salt stress. More specifically, the seed germination rate, root length, chlorophyll content and proline content were significantly higher in transgenic plants ectopically expressing FtWRKY46 than in WT plants after salt stress (P < 0.05), while MDA levels were significantly lower than in WT plants (P < 0.05). Additionally, salt treatment increased the expression of stress-related genes. To summarize, our results suggest that ectopic expression of FtWRKY46 enhance the stress tolerance of transgenic plants by modulating ROS clearance and stress-related gene expression.

Genome-wide investigation of the heat shock transcription factor (Hsf) gene family in Tartary buckwheat (Fagopyrum tataricum).

BACKGROUND: Heat shock transcription factor (Hsfs) is widely found in eukaryotes and prokaryotes. Hsfs can not only help organisms resist high temperature, but also participate in the regulation of plant growth and development (such as involved in the regulation of seed maturity and affects the root length of plants). The Hsf gene was first isolated from yeast and then gradually found in plants and sequenced, such as Arabidopsis thaliana, rice, maize. Tartary buckwheat is a rutin-rich crop, and its nutritional value and medicinal value are receiving more and more attention. However, there are few studies on the Hsf genes in Tartary buckwheat. With the whole genome sequence of Tartary buckwheat, we can effectively study the Hsf gene family in Tartary buckwheat. RESULTS: According to the study, 29 Hsf genes of Tartary buckwheat (FtHsf) were identified and renamed according to location of FtHsf genes on chromosome after removing a redundant gene. Therefore, only 29 FtHsf genes truly had the functional characteristics of the FtHsf family. The 29 FtHsf genes were located on 8 chromosomes of Tartary buckwheat, and we found gene duplication events in the FtHsf gene family, which may promote the expansion of the FtHsf gene family. Then, the motif compositions and the evolutionary relationship of FtHsf proteins and the gene structures, cis-acting elements in the promoter, synteny analysis of FtHsf genes were discussed in detail. What's more, we found that the transcription levels of FtHsf in different tissues and fruit development stages were significantly different by quantitative real-time PCR (qRT-PCR), implied that FtHsf may differ in function. CONCLUSIONS: In this study, only 29 Hsf genes were identified in Tartary buckwheat. Meanwhile, we also classified the FtHsf genes, and studied their structure, evolutionary relationship and the expression pattern. This series of studies has certain reference value for the study of the specific functional characteristics of Tartary buckwheat Hsf genes and to improve the yield and quality of Tartary buckwheat in the future.

Tartary buckwheat transcription factor FtbZIP83 improves the drought/salt tolerance of Arabidopsis via an ABA-mediated pathway.

Plants are subjected to a variety of abiotic stresses during their lifetime, and drought and salt stress are some of the main causes of reduced crop yields. Previous studies have shown that AREB/ABFs within bZIP transcription factors are involved in plant drought and salt stress responses in an ABA-dependent manner. However, the properties and functions of AREB/ABFs in Fagopyrum tataricum, a cereal with good resistance to abiotic stresses, are poorly understood. In this study, a gene encoding an AREB/ABF, designated FtbZIP83, was first isolated from Tartary buckwheat. Expression analysis in Tartary buckwheat indicated that FtbZIP83 was significantly induced by abscisic acid (ABA), NaCl and polyethylene glycol (PEG). The overexpression of FtbZIP83 in Arabidopsis resulted in increased drought/salt tolerance, which was attributed not only to higher proline (Pro) contents and antioxidant enzyme activity in transgenic lines compared with controls but also to the lower reactive oxygen species (ROS) accumulation and malondialdehyde (MDA) content. In addition, we found that FtbZIP83 was able to respond to drought and salt stress by upregulating the transcript abundance of downstream ABA-inducible gene. Furthermore, promoter sequence analysis showed that ABREs were present, and the activity of the FtbZIP83 promoter in transgenic Arabidopsis after drought stress was significantly higher than that under normal conditions. Based on the potential signalling pathways involved in AREB/ABFs, we also screened for the interaction protein FtSnRK2.6/2.3, which may phosphorylate FtbZIP83. Collectively, these results provide evidence that FtbZIP83, as a positive regulator, responds to drought/salt stress via an ABA-dependent signalling pathway composed of SnRK2-AREB/ABF.