Flavanone 3-hydroxylase Research Articles

Seasonal variation in phenylpropanoid biosynthesis and in vitro antioxidant activity of Sorbus domestica leaves: Harvesting time optimisation for medicinal application

Sorbus domestica L. leaves are a phenylpropanoid-rich herbal medicine with antioxidant and anti-inflammatory activity. Here, to optimise harvesting time of the leaves for medicinal application, seasonal variability in the expression of genes encoding enzymes involved in phenylpropanoid biosynthesis (qRT-PCR, quantitative real time polymerase chain reaction) and its relationship with changes in the phytochemical profile (UHPLC-PDA-ESI-MS3, HPLC-PDA-fingerprint, spectrophotometric methods) and antioxidant properties (2,2-diphenyl-1-picrylhydrazyl scavenging and ferric reducing antioxidant power) was evaluated during foliar development. Significant quantitative differences in polyphenolic composition were revealed between spring leaves (predominance of phenylpropanoid acids and flavonoids) and those harvested during the rest of the growing season (predominance of proanthocyanidins, major active components of S. domestica), while their qualitative composition was stable. A set of 40 compounds were detected in the samples by LC–MS/MS including 38 phenylpropanoids. The total phenylpropanoid level (HPLC) in dried leaves dropped from 25.7 mg/g in May to 20.7 mg/g in October but proanthocyanidins and the antioxidant capacity increased simultaneously up to 12.7 mg/g and 49.7 mmol Trolox equivalents/100 g, respectively. Based on the correlation between gene expression and total phenolics and proanthocyanidins (r > 0.825, p < 0.05), phenylalanine ammonia lyase (PAL) and 4-coumaroyl:CoA ligase (4CL) appeared to be core regulators of the phenolic profile among analysed genes (PAL; 4CL; C4H, cinnamate 4-hydroxylase; CHS, chalcone synthase; F3H, flavanone 3-hydroxylase; FLS, flavonols synthase). Due to the synergy of individual components, variability in activity parameters was relatively moderate. Considering the phenylpropanoid levels and antioxidant capacity, the late summer and autumn months (August-October) might be the most advantageous time for harvesting leaf material of optimal quality.

Transcriptome analysis of Salix matsudana under cadmium stress

With the expanded application of heavy metal cadmium, soil cadmium pollution is more and more serious. In this study, using Salix matsudana as a phytoremediation candidate, we observed changes of gene expression and metabolic pathway after 1, 7 and 30 days under 2.5 mg/L and 50 mg/L cadmium stress. The result of transcriptome sequencing showed that we obtained 102 595 Unigenes; 26 623 and 32 154 differentially expressed genes (DEG) in the same concentration and different stress time; 8 550, 3 444 and 11 428 DEG with different concentrations at the same time; 25 genes closely related to cadmium stress response were screened. The changes of genes expression (such as metallothionein, ABC transporter, zinc and manganese transporter) depended on both concentration of cadmium and exposure time. The expression of several genes was obviously up-regulated after cadmium stress, for example 3,6-deoxyinosinone ketolase (ROT3) in brassinolide synthesis pathway and flavonoid synthase (FLS), flavanone-3-hydroxylase (F3H) in the synthesis pathway of brassinolide. In addition, GO analysis shows that GO entries were mainly enriched in metabolic processes including cellular processes, membranes, membrane fractions, cells, cellular fractions, catalytic activation and binding proteins in response to cadmium stress, whose number would increase along with cadmium concentration and exposure time. The reliability of transcriptome information was verified by qPCR and physiological experimental data. Response mechanisms of S. matsudana after cadmium stress were analyzed by transcriptome sequencing, which provided theoretical guidance for remediation of cadmium pollution in soil by S. matsudana.

The Effect of Ethylene on the Color Change and Resistance to Botrytis cinerea Infection in 'Kyoho' Grape Fruits.

The formation of grape quality and the mechanism of resistance against foreign pathogens affect the storage stability of fruits during post-harvest handling. Ethylene plays a crucial role in regulating the ripeness of fruits and can be used as an exogenous regulator to resist exogenous pathogens. In this study, we used different concentrations of ethephon for treatment of grape fruits before veraison, analyzed the anthocyanin content, soluble solids, titratable acid, and determined fruit firmness and cell wall metabolism-related enzymes during fruit development. Results showed that exogenous ethephon promoted the early coloration of grape fruits and increased the coloring-related genes myeloblastosis A1(MYBA1), myeloblastosis A2(MYBA2), chalcone isomerase (CHI), flavanone 3-hydroxylase (F3H), flavonoid 3’-hydroxylase gene (F3’H), flavonoid 3’, 5’hydroxylase (F3’5’H), 3-O-flavonoid glucosyltransferase (UFGT), and glutathione S-transferase (GST), softening related genes Polygalacturonase(PG), pectinate lyases(PL) and Pectin methylesterase( PME, as well as ethylene metabolism pathway-related genes 1-aminocyclopropane-1-carboxylic acid synthase 1(ACS1), 1-aminocyclopropane-1-carboxylic acid oxidase 2 (ACO2), ethylene receptor gene(ETR2), and ethylene-insensitive 3 (EIN3). Ethephon treatment also increased soluble solids and decreased titratable acid in grape fruit. Fruits pretreated with ethephon were inoculated with Botrytis cinerea, which led to resistance in grape fruit through activation of the antioxidant system. The expression levels of disease resistance-related genes including VvPAD4, VvPIP1, VvNAC26, VvDREB, VvAPX, Vvpgip, VvWRKY70, VvMYC2, VvNPR1 also increased in inoculated fruit with pathogen following ethephon pretreatment. Furthermore, we monitored ethylene response factor 1(ERF1) transcription factor, which could interact with protein EIN3 during ethylene signal transduction and mediate fruit resistance against B. cinerea infection. Meanwhile, overexpression of VvERF1 vectorin strawberry fruits reduced the susceptibility to B. cinerea infection. We suggest that ethylene can induce resistance in ripened fruits after B. cinerea infection and provide adequate postharvest care.

Transcriptome Sequencing and Chemical Analysis Reveal the Formation Mechanism of White Florets in Carthamus tinctorius L.

Carthamus tinctorius L. (safflower), an economic crop and herb, has been extensively studied for its diverse chemical constituents and pharmacological effects, but the mechanism of safflower pigments (SP) leading to different colors of florets has not been clarified. In the present study, we compared the contents of SP in two varieties of safflower with white and red florets, named Xinhonghua No. 7 (WXHH) and Yunhong No. 2 (RYH). The results showed the contents of SP in RYH were higher than WXHH. To investigate genes related to SP, we obtained six cDNA libraries of florets from the two varieties by transcriptome sequencing. A total of 225,008 unigenes were assembled and 40 unigenes related to safflower pigment biosynthesis were annotated, including 7 unigenes of phenylalanine ammonia-lyase (PAL), 20 unigenes of 4-coumarate-CoA ligase (4CL), 1 unigene of trans-cinnamate 4-monooxygenase (C4H), 7 unigenes of chalcone synthase (CHS), 4 unigenes of chalcone isomerase (CHI), and 1 unigene of flavanone 3-hydroxylase (F3H). Based on expression levels we selected 16 differentially expressed unigenes (DEGs) and tested them using reverse transcription-quantitative real-time polymerase chain reaction (RT-qPCR), which was consistent with the sequencing results. Consequently, we speculated that in WXHH, 3 PALs, 3 4CLs, 1 C4H, 1 CHS, and 1 CHI, which were down-regulated, and 1 F3H, which was up-regulated, may play a key role in the formation of white florets.

UV-B radiation hormesis in broccoli florets: Glucosinolates and hydroxy-cinnamates are enhanced by UV-B in florets during storage

Abstract Abiotic stresses are oxidative in nature and cause generation of reactive oxygen species (ROS) in plant bodies. Severe stresses can be harmful to the plant tissue, whereas sub-acute or lower doses of stresses could enhance or induce protective mechanisms, a biological phenomenon known as hormesis. The objective of this work was to examine the effect of hormetic as well as high doses of UV-B on the quality along with glucosinolate and hydroxy-cinnamate contents in broccoli florets during storage. An UV-B dose of 1.5 kJ m-2 was found to be hormetic from the color retention response. Color development, weight loss and respiration rate were monitored during 21 d of storage at 4 °C. The gene expression of dihomomethionine N-hydroxylase (CYP79F1), tryptophan N-hydroxylase 2 (CYP79B3), phenylalanine N-hydroxylase (CYP79A2), phenylalanine ammonia-lyase (PAL), chalcone synthase (CH) and flavanone 3-hydroxylase (F3H1) in the treated broccoli was also evaluated. The antioxidant capacity and the profiles of glucosinolates and hydroxy-cinnamates were determined for up to 14 d in broccoli florets stored at 4 °C by LC–MS. The hormetic dose of UV-B was effective in delaying the yellowing of broccoli florets. The initial respiration rate of the florets treated with the hormetic and a high dose (7.2 kJ m-2) was significantly high. The antioxidant capacity of florets was higher in UV-B treated florets relative to the control. The titers of indole-type glucosinolates and hydroxycinnamates in broccoli were significantly (p

Comparative metabolomic analyses of Dendrobium officinale Kimura et Migo responding to UV-B radiation reveal variations in the metabolisms associated with its bioactive ingredients.

BackgroundDendrobium officinale Kimura et Migo, a member of the genus Dendrobium, is a traditional Chinese medicine with high commercial value. The positive roles of UV-B radiation on active ingredient metabolism in various medicinal plants have been studied. However, the metabolic responses of D. officinale stems to UV-B treatment is largely unknown.MethodsAn untargeted metabolomics method was used to investigate the metabolic variations in D. officinale stems between the control and UV-B treatments.ResultsIn total, 3,655 annotated metabolites, including 640 up- and 783 down-regulated metabolites, were identified and grouped into various primary metabolic categories. Then, a number of metabolites involved in the polysaccharide, alkaloid and flavonoid biosynthesis pathways were identified. For polysaccharide biosynthesis, several intermediate products, such as pyruvate, secologanate, tryptophan and secologanin, were significantly up-regulated by the UV-B treatment. For polysaccharide biosynthesis, many key fundamental building blocks, from the glycolysis, starch and sucrose metabolism, and fructose and mannose metabolism pathways, were induced by the UV-B treatment. For flavonoid metabolism, accumulations of several intermediate products of chalcone synthase, chalcone isomerase and flavanone 3-hydroxylase were affected by the UV-B treatment, indicating an involvement of UV-B in flavonoid biosynthesis. The UV-B induced accumulation of polysaccharides, alkaloids and flavonoids was confirmed by HPLC analysis. Our study will help to understand the effects of UV-B on the accumulation of active ingredients in D. officinale.

Transgenic expression of flavanone 3‐hydroxylase redirects flavonoid biosynthesis and alleviates anthracnose susceptibility in sorghum

Flavonoids are ubiquitous in terrestrial plants with important physiological functions. The in planta flavonoid profile depends on the activities of different biosynthesis enzymes (Figure 1a). Flavanone 3-hydroxylase (F3H) is a key enzyme channeling carbon flow towards the production of 3-hydroxylated flavonoids, including flavonols and anthocyanidins. In Poaceae, F3H-encoding genes are generally inactive in vegetative tissues which accumulate flavone derivatives as the predominant flavonoid metabolites. Meanwhile, sorghum produces 3-deoxyanthocyanidins and flavones as phytoalexins for defense against pathogens such as Colletotrichum sublineola, the causal agent of anthracnose.

Comparative transcriptome analysis of mulberry reveals anthocyanin biosynthesis mechanisms in black (Morus atropurpurea Roxb.) and white (Morus alba L.) fruit genotypes

BackgroundTo gain a better understanding of anthocyanin biosynthesis in mulberry fruit, we analyzed the transcriptome of the mulberry varieties Da 10 (Morus atropurpurea Roxb., black fruit) and Baisang (Morus alba L., white fruit).ResultsWe found that whereas Da 10 had high levels of cyanidin 3-O-glucoside (Cy), and pelargonidin 3-O-glucoside (Pg), Baisang contained only Cy, at low levels. Based on a comparative transcriptome analysis, we annotated more than 27,085 genes (including 1735 new genes). Genes that were differentially expressed between Da 10 and Baisang were detected at three stages of fruit development: S1 [4256 genes, 10 days post-anthesis (DPA)], S2 (5612 genes, 19 DPA), and S3 (5226 genes, 28 DPA). Anthocyanin biosynthesis was found to be associated with the expression of 15 core genes and 5 transcription factors. Relative to Baisang, Da 10 showed a significant upregulation of genes involved in the early stages (production of the intermediate compounds chalcone and dihydroflavonol) and late stages (production of Cy and Pg) of anthocyanin biosynthesis. Baisang showed a significant downregulation of the genes involved in the early stages of anthocyanin biosynthesis and overexpression of flavanone 3-hydroxylase (FLS), resulting in the generation of quercetin and/or myricetin but not anthocyanins.ConclusionsThe biosynthesis of anthocyanin in mulberry fruit is initiated from the precursor, phenylalanine, and mediated by the upregulation of dihydroflavonol 4-reductase, anthocyanidin synthase, anthocyanidin 3-O-glucosyltransferase, and cyanidin-3-O-glucoside 2-O-glucuronosyltransferase, and downregulation of FLS to produce Cy and Pg.

Genome-wide variations analysis of sorghum cultivar Hongyingzi for brewing Moutai liquor

BackgroundHongyingzi is a sorghum (Sorghum bicolor L. Moench) cultivar for brewing Moutai liquor. For an overall understanding of the whole genome of Hongyingzi, we performed whole-genome resequencing technology to reveal its comprehensive variations.ResultsCompared with the BTx623 reference genome, we uncovered 1,885,774 single nucleotide polymorphisms (SNPs), 309,381 small fragments insertions and deletions (Indels), 31,966 structural variations (SVs), and 217,273 copy number variations (CNVs). These alterations conferred 29,614 gene variations. It was also predicted that 35 gene variations were related to the multidrug and toxic efflux (MATE) transporter, chalcone synthase (CHS), ATPase isoform 10 (AHA10) transporter, dihydroflavonol-4-reductase (DFR), the laccase 15 (LAC15), flavonol 3′-hydroxylase (F3′H), flavanone 3-hydroxylase (F3H), O-methyltransferase (OMT), flavonoid 3′5′ hydroxylase (F3′5′H), UDP-glucose:sterol-glucosyltransferase (SGT), flavonol synthase (FLS), and chalcone isomerase (CHI) involved in the tannin synthesis.ConclusionsThese results would provide theoretical supports for the molecular markers developments and gene function studies related to the tannin synthesis, and the genetic improvement of liquor-making sorghum based on the genome editing technology.

Flavonoids are indispensable for complete male fertility in rice.

Flavonoids are essential for male fertility in some but not all plant species. In rice (Oryza sativa), the chalcone synthase mutant oschs1 produces flavonoid-depleted pollen and is male sterile. The mutant pollen grains are viable with normal structure, but they display reduced germination rate and pollen-tube length. Analysis of oschs1/+ heterozygous lines shows that pollen flavonoid deposition is a paternal effect and fertility is independent of the haploid genotypes (OsCHS1 or oschs1). To understand which classes of flavonoids are involved in male fertility, we conducted detailed analysis of rice mutants for branch-point enzymes of the downstream flavonoid pathways, including flavanone 3-hydroxylase (OsF3H; flavonol pathway entry enzyme), flavone synthase II (CYP93G1; flavone pathway entry enzyme), and flavanone 2-hydroxylase (CYP93G2; flavone C-glycoside pathway entry enzyme). Rice osf3h and cyp93g1 cyp93g2 CRISPR/Cas9 mutants, and cyp93g1 and cyp93g2 T-DNA insertion mutants showed altered flavonoid profiles in anthers, but only the osf3h and cyp93g1 cyp93g2 mutants displayed reduction in seed yield. Our findings indicate that flavonoids are essential for complete male fertility in rice and a combination of different classes (flavanones, flavonols, flavones, and flavone C-glycosides) appears to be important, as opposed to the essential role played primarily by flavonols that has been previously reported in several plant species.

Expression profile of genes involved in ramie flavonoids biosynthesis pathway and regulation of flavanone 3-hydroxylase (BnF3H) in response to aquatic environment

Flavonoids contained in ramie (Boehmeria nivea) root are important resources for officinal use. The plant seedling can grow optimally and develop an easy-harvesting adventitious root system in a hydroponic system. This research was carried out to unravel the regulation mechanism of flavonoid biosynthesis under hydroponic culturing condition, which hitherto remained unknown. Root samples of the ramie plants grown in soil and hydroponic conditions at different developmental stages were collected and pooled for transcriptome sequencing, and the expression levels of flavonoid biosynthesis-related genes and total flavonoid content were quantified. A total of 43,541 unigenes were finally annotated and the expression of 9 key unigenes were confirmed by qRT-PCR. The flavonoid biosynthesis-related gene BnF3H was significantly up-regulated in the roots of hydroponic ramie, and higher total flavonoids content in the hydroponic ramie roots was also observed. The large dataset of transcripts and unigenes provided abundant genetic information for identifying flavonoid biosynthesis genes in ramie roots. Our results suggest flavonoid biosynthesis in ramie roots is positively affected in aquatic environment, and hydroponic culturing could have a potential for enhancing total flavonoids content in ramie roots.

Characterization and expression analysis of four members genes of flavanone 3-hydroxylase families from Chamaemelum nobile

Chamaemelum nobile is a traditional Chinese herbal medicine, whose secondary metabolites used in the pharmacology of Chinese medicine. Among them, the flavonoids have great research value. Flavanone 3-hydroxylase (F3H) is one of the core enzymes in the early steps of flavonoid biosynthesis. This study aimed to elucidate the structures, functions, and expression levels of F3H families from C. nobile. Four members of the F3H family were screened from C. nobile transcriptome data and performed bioinformatics analysis. Results showed that CnF3H1~4 had a high similarity with the other F3H plants, and all genes contained two conserved isopenicillin N synthase-like and oxoglutarate/iron-dependent dioxygenase domains. Further analysis revealed that the four CnF3H proteins contained some differences in binding sites. The results of secondary and 3-D structures displayed that the composition and proportion of the four CnF3H secondary structures were basically the same, and their 3D structures were consistent with the secondary structures. The phylogenetic tree displayed that CnF3H2, CnF3H3, and CnF3H4 were grouped with Asteraceae. The expression patterns of CnF3Hs in the roots, stems, leaves, and flowers of C. nobile were evaluated using the value of RPKM. The results indicated that CnF3Hs had significant difference in the expression of different tissues. Especially, CnF3H1~3 and CnF3H4 had the highest expression levels in the flowers and roots, respectively. Hence, CnF3Hs played a significant role in the flavonoid metabolism.

CcMYB6-1 and CcbHLH1, two novel transcription factors synergistically involved in regulating anthocyanin biosynthesis in cornflower

Anthocyanins in cornflower (Centaurea cyanus) is catalysed by a set of biosynthesis genes, however, the potential mechanism of transcriptional regulation remains unclear. In the present study, we traced the dynamic changes of petal colour development from white to violet and finally to blue on the same petal in cornflower. Pigment analysis showed that anthocyanin accumulation dramatically increased with petal colour development. Subsequently, nine libraries from above three colour regions were constructed for RNA-seq and 105,506 unigenes were obtained by de novo assembling. The differentially expressed genes among three colour regions were significantly enriched in the phenylpropanoid biosynthesis and flavonoid biosynthesis pathways, leading to the excavation and analysis of 46 biosynthesis genes involved in this process. Furthermore, four R2R3-CcMYBs clustered into subgroup 4 or subgroup 6 and one CcbHLH1 clustered into IIIf subgroup were screened out by phylogenetic analysis with Arabidopsis homologues. The promoters of flavanone 3-hydroxylase (CcF3H) and dihydroflavonol 4-reductase (CcDFR) were further isolated to investigate upstream regulation mechanism. CcMYB6-1 significantly upregulated the activity of above two promoters and stimulated anthocyanin accumulation by dual luciferase assay and transient expression in tobacco leaves, and its activity was obviously enhanced when co-infiltrated with CcbHLH1. Moreover, both yeast two-hybrid and bimolecular fluorescence complementation assays indicated the protein-protein interaction between these two activators. Based on these obtained results, it reveals that CcMYB6-1 and CcbHLH1 are two novel transcription factors synergistically involved in regulating anthocyanin biosynthesis. This study provides insights into the regulatory mechanism of anthocyanin accumulation in cornflower.

Coexpression of PalbHLH1 and PalMYB90 Genes From Populus alba Enhances Pathogen Resistance in Poplar by Increasing the Flavonoid Content.

Secondary metabolites of the flavonoid pathway participate in plant defense, and bHLH and MYB transcription factors regulate the synthesis of these metabolites. Here, we define the regulatory mechanisms in response to pathogens. Two transcription factors from Populus alba var. pyramidalis, PalbHLH1 and PalMYB90, were overexpressed together in poplar, and transcriptome analysis revealed differences in response to pathogen infection. The transgenic plants showed elevated levels of several key flavonoid pathway components: total phenols, proanthocyanidins (PAs), and anthocyanins and intermediates quercetin and kaempferol. Furthermore, PalbHLH1 and PalMYB90 overexpression in poplar enhanced antioxidase activities and H2O2 release and also increased resistance to Botrytis cinerea and Dothiorella gregaria infection. Gene expression profile analysis showed most genes involved in the flavonoid biosynthesis pathway or antioxidant response to be upregulated in MYB90/bHLH1-OE poplar, but significant differential expression occurred in response to pathogen infection. Specifically, expression of PalF3H (flavanone 3-hydroxylase), PalDFR (dihydroflavonol 4-seductase), PalANS (anthocyanin synthase), and PalANR (anthocyanin reductase), which function in initial, middle, and final steps of anthocyanin and PA biosynthesis, respectively, was significantly upregulated in D. gregaria-infected MYB90/bHLH1-OE poplar. Our results highlight that PalbHLH1 and PalMYB90 function as transcriptional activators of flavonoid pathway secondary-metabolite synthesis genes, with differential mechanisms in response to bacterial or fungal infection.

Increased Flavonol Levels in Tobacco Expressing AcFLS Affect Flower Color and Root Growth

The onion (Allium cepa L.) flavonol synthase (AcFLS-HRB) gene, encoding an enzyme responsible for flavonol biosynthesis in yellow onion, was recently identified and enzymatically characterized. Here, we performed an in vivo feeding assay involving bacterial expression of AcFLS-HRB and observed that it exhibited both flavanone 3-hydroxylase (F3H) and FLS activity. Transgenic tobacco (Nicotiana tabacum) expressing AcFLS-HRB produced lighter-pink flowers compared to wild-type plants. In transgenic petals, AcFLS-HRB was highly expressed at the mRNA and protein levels, and most AcFLS-HRB protein accumulated in the insoluble microsomal fractions. High-performance liquid chromatography (HPLC) analysis showed that flavonol levels increased but anthocyanin levels decreased in transgenic petals, indicating that AcFLS-HRB is a functional gene in planta. Gene expression analysis showed the reduced transcript levels of general phenylpropanoid biosynthetic genes and flavonoid biosynthetic genes in AcFLS-HRB overexpressed tobacco petals. Additionally, transgenic tobacco plants at the seedling stages showed increased primary root and root hair length and enhanced quercetin signals in roots. Exogenous supplementation with quercetin 3-O-rutinoside (rutin) led to the same phenotypic changes in root growth, suggesting that rutin is the causal compound that promotes root growth in tobacco. Therefore, augmenting flavonol levels affects both flower color and root growth in tobacco.

The Effects of Ultraviolet A/B Treatments on Anthocyanin Accumulation and Gene Expression in Dark-Purple Tea Cultivar 'Ziyan' (Camellia sinensis).

‘Ziyan’ is a novel anthocyanin-rich tea cultivar with dark purple young shoots. However, how its anthocyanin accumulation is affected by environmental factors, such as ultraviolet (UV), remains unclear. In this study, we observed that UV light treatments stimulated anthocyanin accumulation in ‘Ziyan’ leaves, and we further analyzed the underlying mechanisms at gene expression and enzyme activity levels. In addition, the catechins and chlorophyll contents of young shoots under different light treatments were also changed. The results showed that the contents of total anthocyanins and three major anthocyanin molecules, i.e., delphinidin, cyanidin, and pelargonidin, were significantly higher in leaves under UV-A, UV-B, and UV-AB treatments than those under white light treatment alone. However, the total catechins and chlorophyll contents in these purple tea plant leaves displayed the opposite trends. The anthocyanin content was the highest under UV-A treatment, which was higher by about 66% than control. Compared with the white light treatment alone, the enzyme activities of chalcone synthase (CHS), flavonoid 3′,5′-hydroxylase (F3′5′H), and anthocyanidin synthase (ANS) under UV treatments increased significantly, whereas the leucoanthocyanidin reductase (LAR) and anthocyanidin reductase (ANR) activities reduced. There was no significant difference in dihydroflavonol 4-reductase (DFR) activity under all treatments. Comparative transcriptome analyses unveiled that there were 565 differentially expressed genes (DEGs) of 29,648 genes in three pair-wise comparisons (white light versus UV-A, W vs. UV-A; white light versus UV-B, W vs. UV-A; white light versus UV-AB, W vs. UV-AB). The structural genes in anthocyanin pathway such as flavanone 3-hydroxylase (F3H), F3′5′H, DFR, and ANS, and regulatory gene TT8 were upregulated under UV-A treatment; F3′5′H, DFR, ANS, and UFGT and regulatory genes EGL1 and TT2 were upregulated under UV-AB treatment. However, most structural genes involved in phenylpropanoid and flavonoid pathways were downregulated under UV-B treatment compared with control. The expression of LAR and ANR were repressed in all UV treatments. Our results indicated that UV-A and UV-B radiations can induce anthocyanin accumulation in tea plant ‘Ziyan’ by upregulating the structural and regulatory genes involved in anthocyanin biosynthesis. In addition, UV radiation repressed the expression levels of LAR, ANR, and FLS, resulting in reduced ANR activity and a metabolic flux shift toward anthocyanin biosynthesis.

A Transcriptional Network Makes Normal Tomato Fruit Not Purple

A Transcriptional Network Makes Normal Tomato Fruit Not Purple

Transcriptome sequencing flower petals reveals insights into regulation of flavonoid biosynthesis in Osmanthus fragrans

Osmanthus fragrans Lour., one of the top 10 most popular flowers in China, is known for both its beauty and fragrance. It is rich in flavonoids, a class of secondary metabolites with significant neuroprotective, free-radical scavenging, and anti-oxidant activity. To understand the mechanisms regulating flavonoid biosynthesis, we conducted transcriptome sequencing O. fragrans flowers to analyze gene expressions during the full flowering stage. The RNA was isolated separately from petals of cvs. Yingui and Dangui, which were treated or not with jasmonic acid, salicylic acid, or abscisic acid. A total of 142 029 unigenes were denovo assembled, and 50 918 unigenes were annotated. The differentially expressed genes were identified, annotated, and classified. The results of transcriptome sequencing and real-time PCR revealed higher expressions of phenylalanine ammonia-lyase (PAL), PAL1, chalcone synthase (CHS), flavanone-3-hydroxylase (F3H), flavonol synthase (FLS), and lower expressions of dihydroflavonol-4-reductase (DFR), anthocyanidin synthase (ANS) in 'Yingui' than in 'Dangui'. Such an expression pattern facilitated the higher accumulation of flavonoids in 'Yingui'. Several genes of the flavonoid biosynthesis pathway were upregulated by jasmonic acid and salicylic acid in both the cultivars leading to flavonoid accumulation in their petals. In the v-myb avian myeloblastosis viral oncogene homolog 1(MYB1)-overexpressing petals, the expressions of PAL, PAL1, CHI, and FLS increased. The results suggest that MYB1 may participate in the flavonoid biosynthesis pathway and regulate the expression of some upstream genes in O. fragrans.

Multiplex CRISPR/Cas9-mediated metabolic engineering increases soya bean isoflavone content and resistance to soya bean mosaic virus.

SummaryIsoflavonoids, which include a variety of secondary metabolites, are derived from the phenylpropanoid pathway and are distributed predominantly in leguminous plants. These compounds play a critical role in plant–environment interactions and are beneficial to human health. Isoflavone synthase (IFS) is a key enzyme in isoflavonoid synthesis and shares a common substrate with flavanone‐3‐hydroxylase (F3H) and flavone synthase II (FNS II). In this study, CRISPR/Cas9‐mediated multiplex gene‐editing technology was employed to simultaneously target GmF3H1, GmF3H2 and GmFNSII‐1 in soya bean hairy roots and plants. Various mutation types and frequencies were observed in hairy roots. Higher mutation efficiencies were found in the T0 transgenic plants, with a triple gene mutation efficiency of 44.44%, and these results of targeted mutagenesis were stably inherited in the progeny. Metabolomic analysis of T0 triple‐mutants leaves revealed significant improvement in isoflavone content. Compared with the wild type, the T3 generation homozygous triple mutants had approximately twice the leaf isoflavone content, and the soya bean mosaic virus (SMV) coat protein content was significantly reduced by one‐third after infection with strain SC7, suggesting that increased isoflavone content enhanced the leaf resistance to SMV. The isoflavone content in the seeds of T2 triple mutants was also significantly increased. This study provides not only materials for the improvement of soya bean isoflavone content and resistance to SMV but also a simple system to generate multiplex mutations in soya bean, which may be beneficial for further breeding and metabolic engineering.

Identification of the Genes Involved in Anthocyanin Biosynthesis and Accumulation in Taxus chinensis.

Taxus chinensis is a precious woody species with significant economic value. Anthocyanin as flavonoid derivatives plays a crucial role in plant biology and human health. However, the genes involved in anthocyanin biosynthesis have not been identified in T. chinensis. In this study, twenty-five genes involved in anthocyanin biosynthesis were identified, including chalcone synthase, chalcone isomerase, flavanone 3-hydroxylase, anthocyanidin synthase, flavonoid 3’-hydroxylase, flavonoid 3’,5’-hydroxylase, dihydroflavonol 4-reductase, anthocyanidin reductase, and leucoanthocyanidin reductase. The conserved domains and phylogenetic relationships of these genes were characterized. The expression levels of these genes in different tissues and different ages of xylem were investigated. Additionally, the anthocyanin accumulation in xylem of different ages of T. chinensis was measured. The results showed the anthocyanin accumulation was correlated with the expression levels of dihydroflavonol 4-reductase, anthocyanidin synthase, flavonoid 3’-hydroxylase, and flavonoid 3’,5’-hydroxylase. Our results provide a basis for studying the regulation of the biosynthetic pathway for anthocyanins and wood color formation in T. chinensis.