Chalcone Isomerase Research Articles

Proanthocyanidin biosynthesis in the developing wheat seed coat investigated by chemical and RNA-Seq analysis.

The composition of proanthocyanidins in the testa (seed coat) of bread wheat was analyzed by thiolysis of PA oligomers from developing grain and found to consist of (+)‐catechin monomers, with a small amount of (+)‐gallocatechin. The average chain length of soluble PA stayed relatively constant between 10 and 20 days post‐anthesis, whereas that of unextractable PA increased over the same period, suggesting that increases in chain length might account for the insolubility of PAs from mature wheat grain. We carried out RNA‐Seq followed by differential expression analysis from dissected tissues of developing grain from red‐ and white‐grained near‐isogenic lines differing in the presence of an active R gene that encodes a MYB transcription factor involved in control of PA biosynthesis. In addition to genes already identified encoding chalcone synthase, chalcone isomerase, flavanone 3‐hydroxylase, and dihydroxyflavonoid 4‐reductase, we showed that wheat genes encoding phenylalanine ammonia lyase, flavonoid 3′,5′‐hydroxylase, leucoanthocyanidin reductase, and a glutathione S‐transferase (the orthologue of maize Bronze‐2) were more highly expressed in the red NIL. We also identified candidate orthologues of other catalytic and regulatory components of flavonoid biosynthesis in wheat.

MsMYB741 is involved in alfalfa resistance to aluminum stress by regulating flavonoid biosynthesis.

Aluminum (Al) toxicity severely restricts plant growth in acidic soils (pH < 5.0). In this study, an R2R3-MYB transcription factor (TF) gene, MsMYB741, was cloned from alfalfa. Its function and gene regulatory pathways were studied via overexpression and RNA interference of MsMYB741 in alfalfa seedlings. Results showed that root elongation increased as a result of MsMYB741 overexpression (MsMYB741-OE) and decreased with MsMYB741 RNA interference (MsMYB741-RNAi) in alfalfa seedlings compared with the wild-type under Al stress. These were attributed to the reduced Al content in MsMYB741-OE lines, and increased Al content in MsMYB741-RNAi lines. MsMYB741 positively activated the expression of phenylalanine ammonia-lyase 1 (MsPAL1) and chalcone isomerase (MsCHI) by binding to MYB and ABRE elements in their promoters, respectively, which directly affected flavonoid accumulation in roots and secretion from root tips in plants under Al stress, eventually affecting Al accumulation in alfalfa. Additionally, MsABF2 TF directly activated the expression of MsMYB741 by binding to the ABRE element in its promoter. Taken together, our results indicate that MsMYB741 transcriptionally activates MsPAL1 and MsCHI expression to increase flavonoid accumulation in roots and secretion from root tips, leading to increased resistance of alfalfa to Al stress.

Flavonoids metabolism and physiological response to ultraviolet treatments in Tetrastigma hemsleyanum Diels et Gilg.

Tetrastigma hemsleyanum Diels et Gilg is a folk herb in Zhejiang Province with anti-inflammatory, antineoplastic, and anti-oxidation effects. Given its pharmacological activity, T. hemsleyanum is known as New “Zhebawei” and included in the medical insurance system of Zhejiang and other provinces. Flavonoids are the most important components of T. hemsleyanum, and their contents are mainly regulated by ultraviolet (UV) radiation. In this study, the total flavonoid contents, flavonoid monomer contents, and flavonoid synthesis related enzyme activities (phenylalanine ammonia–lyase, chalcone synthase, and chalcone isomerase), anti-oxidant enzyme activities (catalase, peroxidase, and superoxide dismutase), and biochemical indicators (malondialdehyde, free amino acid, soluble protein, and soluble sugar) in the leaves (L) and root tubers (R) of T. hemsleyanum with UV treatments were determined. Three kinds of UV radiation (UV-A, UV-B, and UV-C) and six kinds of radiation durations (15 and 30 min, 1, 2, 3, and 5 h) were used. Appropriate doses of UV-B and UV-C radiation (30 min to 3 h) induced eustress, which contributed to the accumulation of flavonoids and improve protective enzyme system activities and bioactive compound contents. Especially, certain results were observed in several special structures of the flavonoid monomer: quercetin contents in L increased by nearly 20 times, isoquercitrin contents in R increased by nearly 34 times; most of flavonoids with glycoside content, such as quercitrin (19 times), baicalin (16 times), and apigenin-7G (13 times), increased multiple times. Compared with the CK group, the flavonoid synthase activities, anti-oxidant enzyme activities, and biochemical substance contents in L and R all increased with UV treatments. This study provides a theoretical foundation for regulating flavonoids by light factors and improving the quality of T. hemsleyanum in production and medical industries.

Molecular mechanisms of flavonoid accumulation in germinating common bean (Phaseolus vulgaris) under salt stress.

Flavonoids are important secondary metabolites, active biomolecules in germinating beans, and have prominent applications in food and medicine due to their antioxidant effects. Rutin is a plant flavonoid with a wide biological activity range. In this study, flavonoid (rutin) accumulation and its related molecular mechanisms in germinating common bean (Phaseolus vulgaris) were observed at different time points (0–120 h) under salt stress (NaCl). The rutin content increased from germination onset until 96 h, after which a reducing trend was observed. Metabolome analysis showed that salt stress alters flavonoid content by regulating phenylpropanoid (ko00940) and flavonoid (ko00941) biosynthesis pathways, as well as their enzyme activities, including cinnamyl-alcohol dehydrogenase (CAD), peroxidase (POD), chalcone isomerase (CHI), and flavonol synthase (FLS). The RNA-seq and quantitative real-time PCR (qRT-PCR) analyses also showed that these two pathways were linked to changes in flavonoid content following salt treatment. These results reveal that salt stress effectively enhanced rutin content accumulation in germinating beans, hence it could be employed to enhance the functional quality of germinating common beans.

Acetylsalicylic acid and salicylic acid alleviate postharvest leaf senescence in Chinese flowering cabbage (Brassica rapa var. parachinensis)

The leaves of ‘Ningxia Chinese flowering cabbage’ readily become yellow and senesce after harvest. To test the effects of acetylsalicylic acid (ASA) and salicylic acid (SA) on postharvest leafy vegetables, Chinese flowering cabbages were sprayed with 2 mmol L−1 ASA and 1.5 mmol L−1 SA, and stored at 15 °C. Compared with the controls, during storage, ASA- and SA-treated cabbages retained higher levels of total chlorophyll (Chl), Chl a, and Chl b, and lower activities of chlorophyllase, Mg dechelatase, pheophytinase, and Chl-degrading peroxidase, and had more intact chloroplasts. Furthermore, ASA and SA repressed respiration intensity, ethylene rate, the activities of 1-aminocyclopropane-1-carboxylic acid (ACC) synthase and ACC oxidase, and ACC content, decreased reactive oxygen species (ROS) levels, and increased the activities of superoxide dismutase, catalase, ascorbate peroxidase, peroxidase, phenylalanine ammonia lyase, cinnamate 4-hydroxylase, 4-coumarate-CoA ligase, chalcone synthase, chalcone isomerase, and cinnamyl alcohol dehydrogenase, promoting the accumulation of phenolics and flavonoids. Using principal component and hierarchical clustering analyses, leaf senescence was attributed to higher respiration, ethylene synthesis, and ROS levels, while the delay of senescence in treated cabbages resulted from the cooperation of ROS metabolism and the phenylpropanoid pathway. ASA and SA slowed leaf degreening and senescence. Between the two treatments, 2 mmol L−1 ASA was the most effective.

Transcriptomic Analyses Reveal Key Genes Involved in Pigment Biosynthesis Related to Leaf Color Change of Liquidambar formosana Hance.

Liquidambar formosana Hance has a highly ornamental value as an important urban greening tree species with bright and beautiful leaf color. To gain insights into the physiological and molecular mechanisms of L. formosana leaf color change, the leaves of three different clones were sampled every ten days from October 13, 2019, five times in total, which are S1, S2, S3, S4 and S5. Transcriptome sequencing was performed at S1 and S4. The chlorophyll content of the three clones decreased significantly, while the anthocyanins content of the three clones increased significantly in the coloring stage. The anthocyanins content of clone 2 was far more than that of the other two clones throughout the period of leaf color change. The transcriptome analysis showed that six DEGs related to anthocyanins biosynthesis, including CHS (chalcone synthase), CHI (chalcone isomerase), F3′H (flavonoid 3′-hydroxylase), DFR (dihydroflavonol 4-reductase), ANS (anthocyanidin synthase) and FLS (flavonol synthase), were found in three clones. Clone 2 has another three DEGs related to anthocyanins biosynthesis, including PAL (Phenylalanine ammonia-lyase), F3′5′H (flavonoid 3′,5′-hydroxylase) and UFGT (flavonoid 3-O-glucosyltransferase). We lay a foundation for understanding the molecular regulation mechanism of the formation of leaf color by exploring valuable genes, which is helpful for L. formosana breeding.

Biosynthetic mechanisms of isoflavone accumulation affected by different growth patterns in Astragalus mongholicus products

BackgroundAt present, Astragalus mongholicus products on the market represent two growth patterns: imitative wild A. mongholicus (WAM) and cultivated A. mongholicus (CAM). The 6-year-old WAM (A6) and 2-year-old CAM (B2) products are often sold as commodities. This study aimed to explore the effects of the abovementioned growth patterns on the biosynthetic mechanisms of isoflavone accumulation in A. mongholicus products.ResultsIn this paper, the content of calycosin-7-O-β-D-glucoside in 6-year-old WAM (A6) was significantly higher than that in 2-year-old CAM (B2) based on high-performance liquid chromatography. Tissue anatomy indicated that A6 has developed phloem fibers, thickened secondary walls, and a more well-developed vascular system than B2. Thirteen differentially accumulated metabolites were found in A6 and B2 by UHPLC-ESI-Q-TOF-MS/MS, of which isoflavones were highly and significantly enriched in A6. By combining transcriptomics and metabolomics analysis, we found that the metabolomics profile was the same as the transcriptomics profile in both A6 and B2. In total, 11 novel isoflavone-related genes were isolated using BLAST and functional annotation through RNA-Seq and Iso-Seq. The results of integrated analysis, Short Time-series Expression Miner analysis, and Pearson correlation analysis showed that the regulation of four key enzymes, cinnamate 4-hydroxylase, 6-deoxychalcone synthase, chalcone reductase, and chalcone isomerase, led to the high accumulation of isoflavones in A6. In addition, AmUFGT (c778119) and AmUCGT (c303354) were predicted to be 7-O-glycosyltransferases by phylogenetic analysis; these genes catalyze formononetin and calycosin, respectively.ConclusionsThe findings of this work will clarify the differences in the biosynthetic mechanism of isoflavone accumulation between A6 and B2, which will guide the cultivation of A. mongholicus.

Active compound analysis of Ziziphus jujuba cv. Jinsixiaozao in different developmental stages using metabolomic and transcriptomic approaches

Jujube (Ziziphus jujuba Mill.) is a popular fruit with health benefits ascribed to its various metabolites. These metabolites determine the flavors and bioactivities of the fruit, as well as their desirability. However, the dynamics of the metabolite composition and the underlying gene expression that modulate the overall flavor and accumulation of active ingredients during fruit development remain largely unknown. Therefore, we conducted an integrated metabolomic and transcriptomic investigation covering various developmental stages in the jujube cultivar Z. jujuba cv. Jinsixiaozao, which is famous for its nutritional and bioactive properties. A total of 407 metabolites were detected by non-targeted metabolomics. Metabolite accumulation during different jujube developmental stages was examined. Most nucleotides and amino acids and their derivatives accumulated during development, with cAMP increasing notably during ripening. Triterpenes gradually accumulated and were maintained at high concentrations during ripening. Many flavonoids were maintained at relatively high levels in early development, but then rapidly decreased later. Transcriptomic and metabolomic analyses revealed that chalcone synthase (CHS), chalcone isomerase (CHI), flavonol synthase (FLS), and dihydroflavonol 4-reductase (DFR) were mainly responsible for regulating the accumulation of flavonoids. Therefore, the extensive downregulation of these genes was probably responsible for the decreases in flavonoid content during fruit ripening. This study provide an overview of changes of active components in ‘Jinsixiaozao’ during development and ripening. These findings enhance our understanding of flavor formation and will facilitate jujube breeding for improving both nutrition and function.

Efficient production of anthocyanins in Saccharomyces cerevisiae by introducing anthocyanin transporter and knocking out endogenous degrading enzymes

Anthocyanins are natural pigments found in various plants. As multifunctional natural compounds, anthocyanins are widely used in food, pharmaceuticals, health products, and cosmetics. At present, the anthocyanins are heterologously biosynthesized in prokaryotes from flavan-3-ols, which is rather expensive. This study aimed to metabolically engineer Saccharomyces cerevisiae for anthocyanin production. Anthocyanin production has been extensively studied to understand the metabolic pathway enzymes in their natural hosts, including CHS (chalcone synthase); FLS (flavonol synthase); CHI (chalcone isomerase); F3H (flavanone 3-hydroxylase); F3′H (flavonoid 3′-hydroxylase); F3′5′H (flavonoid 3′,5′-hydroxylase); DFR (dihydroflavonol 4-reductase); ANS (anthocyanidin synthase); LAR (leucoanthocyanidin reductase); and UFGT (flavonoid 3-O-glucosyltransferase). The anthocyanin transporter MdGSTF6 was first introduced and proven to be indispensable for the biosynthesis of anthocyanins. By expressing MdGSTF6, FaDFR, PhANS0, and Dc3GT and disrupting EXG1 (the main anthocyanin-degrading enzyme), the BA-22 strain produced 261.6 mg/L (254.5 mg/L cyanidin-3-O-glucoside and 7.1 mg/L delphinidin-3-O-glucoside) anthocyanins from 2.0 g/L dihydroflavonols, which was known to be the highest titer in eukaryotes. Finally, 15.1 mg/L anthocyanins was obtained from glucose by expressing the de novo biosynthesis pathway in S. cerevisiae, which is known to be the highest de novo production. It is the first study to show that through the introduction of a plant anthocyanin transporter and knockout of a yeast endogenous anthocyanin degrading enzyme, the anthocyanin titer has been increased by more than 100 times.

Integrated Transcriptomic and Metabolomic Analysis of the Mechanism of Foliar Application of Hormone-Type Growth Regulator in the Improvement of Grape (Vitis vinifera L.) Coloration in Saline-Alkaline Soil.

(1) Background: To solve the problems of incomplete coloration and quality decline caused by unreasonable use of regulators in grapes, this study clarified the differences in the effects of a hormone-type growth regulator (AUT) and two commercial regulators on grape coloration and quality through field experiments. (2) Methods: The color indexes (brightness (L*), red/green color difference (a*), yellow/blue color difference (b*), and color index for red grapes (CIRG)) of grape fruit were measured using a CR-400 handheld color difference meter. The titratable acid content, total phenol content, and total sugar content were measured using anthrone colorimetry, folinol colorimetry, and NaOH titration, respectively, and the chalcone isomerase activity, phenylalanine ammoniase activity, dihydroflavol reductase activity, and anthocyanin content were measured using a UV spectrophotometer. (3) Results: The a*, total sugar and total phenol contents, and chalcone isomerase (CHI) and phenylalanine ammoniase (PAL) activities of grape fruit in the AUT treatment significantly increased, while the titratable acid content significantly decreased, compared to those in the CK treatment. The expressions of the differentially expressed genes (DEGs) trpB and argJ in AUT treatment were significantly up-regulated. The expressions of the differentially expressed metabolites (DEMs) phenylalanine and 4-oxoproline were significantly up-regulated, while those of 3,4-dihydroxybenzaldehyde and N-acetyl glutamate were significantly down-regulated. The CIRG significantly increased by 36.4% compared to that in the CK, indicating improved fruit coloration. (4) Conclusion: The AUT could shorten the color conversion period of grape fruit and improve the coloration, taste, and tolerance to saline and alkaline stresses.

The spatiotemporal regulations of epicatechin biosynthesis under normal flowering and the continuous inflorescence removal treatment in Fagopyrum dibotrys

BackgroundFlowering is a critical physiological change that interferes with not only biomass yield but also secondary metabolism, such as the biosynthesis of flavonoids, in rhizome/root plants. The continuous inflorescence removal (CIR) treatment is frequently conducted to weaken this effect. Fagopyrum dibotrys (D.Don) H.Hara (Golden buckwheat) is a kind of rhizome medicinal plant rich in flavonoids and is widely used for the treatment of lung diseases. The CIR treatment is usually conducted in F. dibotrys because of its excessive reproductive growth. To uncover the molecular mechanisms, comprehensive analysis was performed using metabolome and transcriptome data obtained from normally bloomed and the CIR treated plants.ResultsMetabolome results demonstrated that in the rhizomes of F. dibotrys, its bioactive compound called epicatechin has higher amount than most of the detected precursors. Compared with the normally bloomed plants, the level of epicatechin in the rhizomes of the CIR group increased by 25% at the withering stage. Based on 96 samples of the control and the CIR groups at 4 flowering stages for 4 tissues, RNA-Seq results revealed a 3 ~ 5 times upregulations of all the key enzyme genes involved in the biosynthesis of epicatechin in both time (from the bud stage to the withering stage) and spatial dimensions (from the top of branch to rhizome) under the CIR treatment compared to normal flowering. Integrated analysis of LC–MS/MS and transcriptome revealed the key roles of several key enzyme genes besides anthocyanidin reductase (ANR). A total of 93 transcription factors were identified to co-expressed with the genes in epicatechin biosynthetic pathway. The flowering activator SQUAMOSA promoter-binding protein like (SPLs) exhibited opposite spatiotemporal expression patterns to that of the epicatechin pathway genes; SPL3 could significantly co-express with all the key enzyme genes rather than the flowering repressor DELLA. Weighted gene co-expression network analysis (WGCNA) further confirmed the correlations among chalcone synthases (CHSs), chalcone isomerases (CHIs), ANRs, SPLs and other transcription factors.ConclusionsSPL3 might dominantly mediate the effect of normal flowering and the CIR treatment on the biosynthesis of epicatechin in rhizomes mainly through the negative regulations of its key enzyme genes including CHS, CHI and ANR.

Response of Anthocyanin Accumulation in Pepper (Capsicum annuum) Fruit to Light Days.

Light is the key factor affecting the synthesis of anthocyanins in pepper. In this study, pepper fruit under different light days was used as experimental material to explore the synthesis of anthocyanins in purple pepper. A total of 38 flavonoid metabolites were identified in the purple pepper germplasm HNUCA21 by liquid chromatography–tandem mass spectrometry (LC-MS/MS), of which 30 belong to anthocyanins. The detected anthocyanin with the highest content was Delphinidin-3-O-glucoside (17.13 µg/g), which reached the maximum after 168 h of light treatment. Through weighted gene co-expression network analysis (WGCNA), the brown module was identified to be related to the early synthesis of anthocyanins. This module contains many structural genes related to flavonoid synthesis, including chalcone synthase (CHS 107871256, 107864266), chalcone isomerase (CHI 107871144, 107852750), dihydroflavonol 4-reductase (DFR 107860031), flavonoid 3′ 5′-hydroxylase (F3’5’H 107848667), flavonoid 3′-monooxygenase (F3M 107862334), leucoanthocyanidin dioxygenase (LDOX 107866341), and trans-cinnamate 4-monooxygenase (TCM 107875406, 107875407). The module also contained some genes related to anthocyanin transport function, such as glutathione S-transferase (GST 107861273), anthocyanidin 3-O-glucosyltransferase (UDPGT 107861697, 107843659), and MATE (107863234, 107844661), as well as some transcription factors, such as EGL1 (107865400), basic helix-loop-helix 104 (bHLH104 107864591), and WRKY44 (107843538, 107843524). The co-expression regulatory network indicated the involvement of CHS, DFR, CHI, and EGL1, as well as two MATE and two WRKY44 genes in anthocyanin synthesis. The identified genes involved in early, middle, and late light response provided a reference for the further analysis of the regulatory mechanism of anthocyanin biosynthesis in pepper.

Mitigating chilling injury of pomegranate fruit skin

Pomegranate cv. Wonderful fruit are susceptible to chilling injury of the peel (CIp) when stored at 7 °C in modified-atmosphere bags for more than 3 months. The damage, manifested as superficial browning, appears first on the fruit skin, i.e., the outer colored layer of the peel. We previously suggested that susceptibility to CIp is associated with downregulation of PHENYLALANINE AMMONIA-LYASE (PAL) in storage, whereas POLYPHENOL OXIDASE (PPO) expression is upregulated. To further explore the role of PPO and PAL in CIp disorder, we monitored their gene expression and enzyme activity in fruit skin collected at three phases of fruit development: premature fruit 7 days before harvest when the fruit is susceptible to CIp disorder; during commercial harvest when fruit skin is aging; and during the following 4 months of storage. In addition, prestorage washing of the fruit with CaCl2 and citric acid solutions, previously shown to reduce CIp incidence in pomegranate and other fruit and to manipulate the PAL-to-PPO ratio, was applied. We determined total phenol content and antioxidative capacity of fruit skin during cold storage, and analyzed the expression of antioxidation-related genes CATALASE, SUPEROXIDE DISMUTASE, and GLUTATHIONE REDUCTASE, and key anthocyanin-biosynthesis genes CHALCONE SYNTHASE, CHALCONE ISOMERASE, and DIHYDROFLAVONOL 4-REDUCTASE. Results suggested that increased skin susceptibility to CIp is related to high total phenol content, relatively high PAL activity/gene expression, and low anthocyanin levels. Although PPO activity and gene expression increased in cold storage, this pattern was observed in all tissues tested, irrespective of their susceptibility to CIp development. Similarly, no association was evident between antioxidation-related genes’ expression levels and CIp susceptibility. On the other hand, pre-storage treatments with CaCl2 and citric acid mainly decreased CIp incidence associated with increased antioxidation-related gene expression. Hence, the mechanism governing mitigation of CIp development by prestorage washing with CaCl2 and citric acid differs from that observed in native CIp-resistant pomegranate skin.

Genome-Wide Classification and Evolutionary Analysis Reveal Diverged Patterns of Chalcone Isomerase in Plants

Flavonoids as a class of important secondary metabolites are widely present in land plants, and chalcone isomerase (CHI) is the key rate-limiting enzyme that participates in catalyzing the stereospecific isomerization of chalcones to yield their corresponding flavanones. However, the phylogenetic dynamics and functional divergence of CHI family genes during the evolutionary path of green plants remains poorly understood. Here, a total of 122 CHI genes were identified by performing a genome-wide survey of 15 representative green plants from the most ancestral basal plant chlorophyte algae to higher angiosperm plants. Phylogenetic, orthologous groups (OG) classification, and genome structure analysis showed that the CHI family genes have evolved into four distinct types (types I–IV) containing eight OGs after gene duplication, and further studies indicated type III CHIs consist of three subfamilies (FAP1, FAP2, and FAP3). The phylogeny showed FAP3 CHIs as an ancestral out-group positioned on the outer layers of the main branch, followed by type IV CHIs, which are placed in an evolutionary intermediate between FAP3 CHIs and bona fide CHIs (including type I and type II). The results imply a potential intrinsic evolutionary connection between CHIs existing in the green plants. The amino acid substitutions occurring in several residues have potentially affected the functional divergence between CHI proteins. This is supported by the analysis of transcriptional divergence and cis-acting element analysis. Evolutionary dynamics analyses revealed that the differences in the total number of CHI family genes in each plant are primarily attributed to the lineage-specific expansion by natural selective forces. The current studies provide a deeper understanding of the phylogenetic relationships and functional diversification of CHI family genes in green plants, which will guide further investigation on molecular characteristics and biological functions of CHIs.

Effects of microwave irradiation of Fagopyrum tataricum seeds on the physicochemical and functional attributes of sprouts

The effects of microwave-preexposing Tartary buckwheat seeds on their germination, including nutritional and functional attributes were investigated. The results showed that microwave irradiation (300 W/50 s) accelerated the germination process. Germination caused a 7.51-fold increase in reducing sugar content and a 69.61% reduction in starch content, accompanied by significantly enhanced amylase activity, which was amplified by 300 W/50 s microwaving. Microwave also enhanced the levels of various amino acids (especially at 300 W/50 s) and fatty acids (especially at 550 W/75 s) during germination. Enhancements of phenylalanine ammonia-lyase (PAL), chalcone isomerase (CHI), and flavonol synthetase (FLS) activities were observed with microwave irradiation, and maximum total flavonoid content (5.15 g/100 g dry weight; 31.78% higher than that of the control) was reached after irradiation at 300 W/50 s. In addition, the antioxidant capacity and the tyrosinase and acetylcholinesterase inhibitory activities were enhanced. Principal component analysis indicated that changes in flavonoids, PAL, CHI, and FLS were highly correlated with antioxidant capacity and enzyme inhibitory activities. Therefore, microwave irradiation is an efficient protocol for enhancing the physicochemical and functional attributes of edible Tartary buckwheat sprouts.

Identification of microRNAs and their target genes related to the accumulation of anthocyanin in purple potato tubers (Solanum tuberosum).

MicroRNAs (miRNAs) are types of endogenous non‐coding small RNAs found in eukaryotes that are 18–25 nucleotides long. miRNAs are considered to be key regulatory factors of the expression of target mRNA. The roles of miRNAs involved in the regulation of anthocyanin accumulation in pigmented potatoes have not been systematically reported. In this study, the differentially expressed miRNAs and their target genes involved in the accumulation of anthocyanin during different developmental stages in purple potato ( Solanum tuberosum L.) were identified using small RNA (sRNA) and degradome sequencing. A total of 275 differentially expressed miRNAs were identified in the sRNA libraries. A total of 69,387,200 raw reads were obtained from three degradome libraries. The anthocyanin responsive miRNA–mRNA modules were analyzed, and 37 miRNAs and 23 target genes were obtained. Different miRNAs regulate the key enzymes of anthocyanin synthesis in purple potato. The structural genes included phenylalanine ammonia lyase, chalcone isomerase, flavanone 3‐hydroxylase, and anthocyanidin 3‐O‐glucosyltransferase. The regulatory genes included WD40, MYB, and SPL9. stu‐miR172e‐5p_L‐1R‐1, stu‐miR828a, stu‐miR29b‐4‐5p, stu‐miR8019‐5p_L‐4R‐3, stu‐miR396b‐5p, stu‐miR5303f_L‐7R + 2, stu‐miR7997a_L‐3, stu‐miR7997b_L‐3, stu‐miR7997c_L + 3R‐5_2ss2TA3AG, stu‐miR156f‐5p_L + 1, stu‐miR156a, stu‐miR156a_R‐1, stu‐miR156e, stu‐miR858, stu‐miR5021, stu‐miR828 and their target genes were validated by qRT‐PCR. They play important roles in the coloration and accumulation of purple potatoes. These results provide new insights into the biosynthesis of anthocyanins in pigmented potatoes.

Transcriptomic analysis identifies differentially expressed genes in purple tender shoots and green mature leaves of Zijuan tea

Purple shoots of tea germplasm Zijuan (Camellia sinensis var. asssamica) have high anthocyanin content and gradually turn green with continued growth and development. To uncover the mechanism of this colour change, we performed transcriptome analysis to identify differentially expressed genes (DEGs). A total of 18 328 DEGs were identified in purple tender shoots of Zijuan (ZJP) compared with mature green leaves of Zijuan (ZJG). These included 85 DEGs associated with phenylpropanoid biosynthesis, 30 DEGs associated with flavonoid biosynthesis, and 3 DEGs involved in flavone and flavonol biosynthesis. We detected higher expression in ZJP than in ZJG of genes encoding the following key enzymes regulating anthocyanin biosynthesis and their corresponding metabolites: chalcone synthase (CHS), anthocyanidin synthase (ANS), anthocyanidin reductase (ANR), adenosine triphosphate (ATP)-binding cassette (ABC) transporters, bifunctional 3-dehydroquinate dehydratase (DHQ), chorismate mutase (CM), cinnamoyl-CoA reductase 1 (CCR), and cinnamyl alcohol dehydrogenase (CAD). In addition, upregulated expression of carbohydrate metabolism, glycolysis pathway, sucrose metabolism, and pyruvate metabolism suggested that glycolysis and upregulation of the sucrose synthase (SUS) gene may provide more intermediates as substrates for promoting anthocyanin accumulation in ZJP. Moreover, DEGs involved in anthocyanin biosynthesis, including phenylalanine ammonialyase (PAL), cinnamate acid 4-hydroxylase (C4H), chalcone isomerase (CHI), flavanone 3-hydroxylase (F3H), flavonol synthase (FLS), ANR, ANS, CHS, and flavonoid 3',5'-hydroxylase (F3'5'H) were significantly (P &amp;lt; 0.01) correlated with the contents of anthocyanin components such as cyanidin (Cy), delphinidin (Dp), peonidin (Pn), petunidin (Pt), and malvidin (Mv). The above results suggested that these upregulated DEGs may contribute to anthocyanin accumulation in ZJP and may play important roles in the colour changes in Zijuan tea. This research provided a foundation for clarifying the mechanisms underlying colour changes in Zijuan tea.

Key genes for phenylpropanoid metabolite biosynthesis during half-highbush blueberry (Vaccinium angustifolium×Vaccinium corymbosum) fruit development

BACKGROUND: Blueberry fruits contain large amounts of phenolic compounds derived from the phenylpropanoid pathway. Their biosynthesis is complex, involving many enzymes. OBJECTIVE: We sought to investigate the content of phenylpropanoid-derived compounds and identify key genes involved in the phenylpropanoid metabolite pathway during half-highbush blueberry fruit development. METHODS: Phenylpropanoid metabolite contents were determined by high-performance liquid chromatography (HPLC) and spectrophotometry. Gene expression was examined through reverse-transcription PCR. RESULTS: Phloretin, chlorogenic acid, total flavonol, quercetin, catechin, and proanthocyanidin contents were highest in small-sized green fruits; myricetin and epicatechin contents were highest in pink fruits; and lignin and anthocyanin were highest in blue fruits. Genes from the 4-coumarate CoA ligase (4CL) family regulate the biosynthesis of phenylpropanoid metabolites. Phenylalanine ammonia-lyase (PAL) and cinnamoyl-CoA reductase (CCR) are key genes in the lignin biosynthetic pathway. Flavonol synthase (FLS) is a key gene affecting total flavonols and the quercetin biosynthetic pathway. PAL and chalcone isomerase (CHI) are key genes in the epicatechin and anthocyanin biosynthetic pathways, respectively. CONCLUSION: Phenylpropanoid metabolites are regulated by multiple genes from the same or different families. Enzymes in different metabolic pathways compete for precursors to form a complex regulatory network for phenylalanine metabolism.

Genetic mapping of sexually dimorphic volatile and non-volatile floral secondary chemistry of a dioecious willow.

Secondary chemistry often differs between sexes in dioecious plant species, a pattern attributed to its possible role in the evolution and/or maintenance of dioecy. We used GC-MS to measure floral volatiles emitted from, and LC-MS to quantitate non-volatile secondary compounds contained in, female and male Salix purpurea willow catkins from an F2 family. Using the abundance of these chemicals, we then performed quantitative trait locus (QTL) mapping to locate them on the genome, identified biosynthetic candidate genes in the QTL intervals, and examined expression patterns of candidate genes using RNA-seq. Male flowers emitted more total terpenoids than females, but females produced more benzenoids. Male tissue contained greater amounts of phenolic glycosides, but females had more chalcones and flavonoids. A flavonoid pigment and a spermidine derivative were found only in males. Male catkins were almost twice the mass of females. Forty-two QTL were mapped for 25 chemical traits and catkin mass across 16 of the 19 S. purpurea chromosomes. Several candidate genes were identified, including a chalcone isomerase associated with seven compounds. A better understanding of the genetic basis of the sexually dimorphic chemistry of a dioecious species may shed light on how chemically mediated ecological interactions may have helped in the evolution and maintenance of dioecy.

Candidate genes for anthocyanin pigmentation in rice stem revealed by GWAS and whole-genome resequencing.

Anthocyanin pigment as a phenolic secondary metabolite is accumulated in areal organs of some rice cultivars. Despite several research attempts, the majority of genomic regions and candidate genes for purple-colored stem (Ps) resulting from anthocyanin pigmentation of rice leaf sheath have not been identified. A genome-wide association study (GWAS) and whole-genome resequencing (WGR) analysis was applied for genetic dissection of anthocyanin pigmentation of rice stem. Using GWAS, the genomic regions (on chromosomes 2, 4, and 6) tagged to eight single-nucleotide polymorphisms (SNPs) were identified to be significantly associated with purple stem, and in the vicinity of GWAS signals, 19 genes were highlighted as putative candidate genes. To narrow down the genomic regions more highly associated to the trait, a WGR study on recombinant inbred lines (RIL) with opposite phenotypes was conducted. After defining the DNA variation between reference genome, maternal parent and the two sister lines, a narrow genomic region on the short arm of chromosome 6 (4.7-6.2 Mbp interval) was identified to be highly associated with anthocyanin pigmentation of rice stem. In the interval, a few candidate genes with probable role in anthocyanin biosynthesis and accumulation were identified, which included five structural genes involved in the known pathways [one chalcone isomerase (CHI), two glycosyl transferases, and two UDP-flavonoid-3-O-glucosyl (UFGT) transferases] and two transcription factors [one basic helix-loop-helix (bHLH)- and one myeloblastosis (MYB)-coding genes]. The identified candidate genes can be used in breeding programs of rice or other Gramineae species for anthocyanin accumulation in areal organs.