Di-π-ethane Rearrangement of Cyano Groups via Energy-Transfer Catalysis.

Molecular rearrangement occupies a pivotal position among fundamental transformations in synthetic chemistry. Radical translocation has emerged as a prevalent synthetic tool, efficiently facilitating the migration of diverse functional groups. In contrast, the development of di-π-methane rearrangement remains limited, particularly in terms of the translocation of cyano functional groups. This is primarily attributed to the energetically unfavorable three-membered-ring transition state. Herein, we introduce an unprecedented di-π-ethane rearrangement enabled by energy-transfer catalysis under visible light conditions. This innovative open-shell rearrangement boasts broad tolerance toward a range of functional groups, encompassing even complex drug and natural product derivatives. Overall, the reported di-π-ethane rearrangement represents a complementary strategy to the development of radical translocation enabled by energy-transfer catalysis.

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.

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.

Diverse biological effects of glycosyltransferase genes from Tartary buckwheat.

BACKGROUND: Tartary buckwheat (Fagopyrum tataricum) is an edible cereal crop whose sprouts have been marketed and commercialized for their higher levels of anti-oxidants, including rutin and anthocyanin. UDP-glucose flavonoid glycosyltransferases (UFGTs) play an important role in the biosynthesis of flavonoids in plants. So far, few studies are available on UFGT genes that may play a role in tartary buckwheat flavonoids biosynthesis. Here, we report on the identification and functional characterization of seven UFGTs from tartary buckwheat that are potentially involved in flavonoid biosynthesis (and have varying effects on plant growth and development when overexpressed in Arabidopsis thaliana.) RESULTS: Phylogenetic analysis indicated that the potential function of the seven FtUFGT proteins, FtUFGT6, FtUFGT7, FtUFGT8, FtUFGT9, FtUFGT15, FtUFGT40, and FtUFGT41, could be divided into three Arabidopsis thaliana functional subgroups that are involved in flavonoid biosynthesis of and anthocyanin accumulation. A significant positive correlation between FtUFGT8 and FtUFGT15 expression and anthocyanin accumulation capacity was observed in the tartary buckwheat seedlings after cold stress. Overexpression in Arabidopsis thaliana showed that FtUFGT8, FtUFGT15, and FtUFGT41 significantly increased the anthocyanin content in transgenic plants. Unexpectedly, overexpression of FtUFGT6, while not leading to enhanced anthocyanin accumulation, significantly enhanced the growth yield of transgenic plants. When wild-type plants have only cotyledons, most of the transgenic plants of FtUFGT6 had grown true leaves. Moreover, the growth speed of the oxFtUFGT6 transgenic plant root was also significantly faster than that of the wild type. At later growth, FtUFGT6 transgenic plants showed larger leaves, earlier twitching times and more tillers than wild type, whereas FtUFGT15 showed opposite results. CONCLUSIONS: Seven FtUFGTs were isolated from tartary buckwheat. FtUFGT8, FtUFGT15, and FtUFGT41 can significantly increase the accumulation of total anthocyanins in transgenic plants. Furthermore, overexpression of FtUFGT6 increased the overall yield of Arabidopsis transgenic plants at all growth stages. However, FtUFGT15 shows the opposite trend at later growth stage and delays the growth speed of plants. These results suggested that the biological function of FtUFGT genes in tartary buckwheat is diverse.

FtMYB8 from Tartary buckwheat inhibits both anthocyanin/Proanthocyanidin accumulation and marginal Trichome initiation.

BACKGROUND: Because flavonoids and trichomes play crucial roles in plant defence, their formation requires fine transcriptional control by multiple transcription factor families. However, little is known regarding the mechanism of the R2R3-MYB transcription factors that regulate both flavonoid metabolism and trichome development. RESULTS: Here, we identified a unique SG4-like-MYB TF from Tartary buckwheat, FtMYB8, which harbours the C2 repression motif and an additional TLLLFR repression motif. The expression profiles of FtMYB8 combined with the transcriptional activity of PFtMYB8 promoter showed that FtMYB8 mRNA mainly accumulated in roots during the true leaf stage and flowering stage and in bud trichomes and flowers, and the expression of this gene was markedly induced by MeJA, ABA and UV-B treatments but repressed by dark treatment. Overexpression of FtMYB8 in Arabidopsis reduces the accumulation of anthocyanin/proanthocyanidin by specifically inhibiting TT12 expression, which may depend on the interaction between FtMYB8 and TT8. Interestingly, this interaction may also negatively regulate the marginal trichome initiation in Arabidopsis leaves. CONCLUSIONS: Taken together, our results suggest that FtMYB8 may fine-tune the accumulation of anthocyanin/proanthocyanidin in the roots and flowers of Tartary buckwheat by balancing the inductive effects of transcriptional activators, and probably regulate trichome distribution in the buds of Tartary buckwheat.

Optimization of mobile phase for the determination of Esomeprazole and related compounds and investigation of stress degradation by LC-MS.

In this study, the objective was to investigate the degradation behavior of Esomeprazole under different recommended stress conditions according to International Conference on Harmonization of Technical Requirements for Registration of Pharmaceuticals for Human Use [1] by HPLC. Our research showed that the effect of mobile phase species on separation was significant for the determination of Esomeprazole and its related compounds. Successful separation of the drug from its related impurities and degradation products formed under different stress conditions was achieved using ammonium acetate buffer/ACN by a gradient elution. Compared with phosphate buffer/ACN, ammonium acetate buffer/ACN under same pH and gradient showed a great improvement in resolution due to the change of elution order. The drug was subjected to stress conditions including acidic, alkaline, oxidative, photolytic, and thermal conditions. Extensive degradation occurred in acidic and oxidative conditions, while mild degradation was observed in alkaline and photolytic conditions. Besides, it turned out the drug was extremely stable under thermal condition. The stability-indicating LC-UV method was validated with respect to linearity, precision, accuracy, specificity, and robustness. The LC-MS method was also adopted for the characterization of degradation products. Based on the m/z values and fragmentation patterns, the degradation pathway of the drug has been proposed.

One-Pot Sequential Hydrogen Isotope Exchange/Reductive Deuteration for the Preparation of α,ß-Deuterated Alcohols using Deuterium Oxide.

An efficient one-pot sequential hydrogen isotope exchange (HIE)/reductive deuteration approach was developed for the preparation of α,ß-deuterated alcohols using ketones as the precursors. The HIE step can also be used for the synthesis of α-deuterated ketones. This method has been applied in the synthesis of four deuterated drug and MS internal standards.

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.

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.

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.