Flavonoid Biosynthetic Pathway Research Articles

A comprehensive study on the bioactive flavonoids in Paulownia flowers: Uncovering metabolic pathways, effective components, and regulatory genes for industrial applications

The study explores Paulownia flowers as an emerging resource in medicine, food, health supplements, cosmetics, and animal feed, despite a lack of characterization of their secondary metabolite accumulation mechanisms. We conducted a comprehensive investigation into the biological activities, metabolite profiles, and gene expression patterns of flavonoids in Paulownia flowers. Our results, obtained using ABTS/FRAP/DPPH assays, MTT assays, and colorimetric analyses, demonstrate the potent antioxidant, antibacterial, hypolipidemic, and antiproliferative activities of Paulownia flower flavonoid (PFF) extracts. Notably, the Paulownia elongata tetraploid (M4) exhibited particularly notable effects. HPLC fingerprinting, molecular docking, and density functional theory calculations led to the identification of rhoifolin and luteolin as key bioactive flavonoids. UV-Vis, fluorescence, and circular dichroism spectroscopy confirmed PFF's role as a reversible and mixed inhibitor of xanthine oxidase (XOD). Transcriptome and metabolomic analyses further revealed substantial enrichment in phenylpropanoid, flavonol, and flavonoid biosynthetic pathways, with PfPP2C, PfPYR, PfPYL, PfNPR1, PfMYBs, and PfbHLHs identified as crucial regulators. This work provides a foundational understanding of flavonoid biosynthesis in Paulownia flowers, paving the way for targeted genetic modification and biotechnological interventions aimed at enhancing flavonoid accumulation for industrial applications.

Genome-wide investigation of family‑1 UDP glycosyltransferases reveals the Fusarium resistance role of VfUGT90A2 in tung trees

UDP-glycosyltransferases (UGTs) play vital roles in pathogen resistance through secondary metabolites, such as flavonoid. Vernicia fordii (tung trees) is well known for its production of industrial oil, which has been decimated by the soil-borne fungus Fusarium oxysporum f. sp. Fordiis (Fof-1). The current understanding of the metabolic and molecular mechanism of V. fordii against Fof-1 is limited. Through genome-wide analysis identified 153 VfUGTs, which were divided into 16 clusters derived from tandem and segmental repeat events. Variations in gene structure and cis-elements have contributed to gene differentiation within the various subfamilies of VfUGTs. VfUGT90A2 was found to be highly induced and expressed, serving as a key gene during the response process to Fof-1 infection in tree roots. The study showed that transgenic hairy roots of the tung trees, overexpressing VfUGT90A2, displayed significantly enhanced resistance to Fof-1. Additionally, the root extract from these transgenic roots was found to have a notable inhibitory effect on the growth of Fof-1 mycelium. Metabolomic and transcriptomic analyses further revealed an increase in flavonoids, such as quercitrin and myricitrin, in the transgenic hairy roots overexpressing VfUGT90A2. Meanwhile, VfUGT90A2 was speculated bound well to quercetin by protein docking predictions. The enzyme activity assay in vitro further verified that VfUGT90A2 catalyzed the glycosylation of quercetin. In all, our data shows that VfUGT90A2 is involved in the flavonoid biosynthetic pathways and has an impact on enhancing the resistance of plant to Fusarium, therefore, offers an opportunity to explore an effective strategy for Fusarium resistant plant breeding.

Foliar flavonoids across an elevation gradient: Plasticity in response to UV, and links with floral pigmentation patterning

Metabolites produced in the flavonoid biosynthetic pathway (FBP) mitigate abiotic stress caused by factors such as ultraviolet (UV) light. Testing whether constitutive flavonoid production or flavonoid plasticity differ between populations spanning ecological gradients can reveal whether geographic patterns are consistent with local adaptation. Abiotic induction of flavonoids can occur in leaves as well as flowers where flavonoids influence UV color patterns perceived by pollinators. Assessing how foliar flavonoids are associated with floral color phenotypes can shed light on how pleiotropy affects biochemical phenotypes across tissues. We exposed Argentina anserina (Rosaceae) plants from alpine and lower elevation populations to low and high levels of UV and measured foliar and petal flavonoid production using UHPLC coupled mass spectrometry. We associated foliar flavonoid abundance with petal flavonoid abundance, and the size of the UV absorbing petal area (‘UV bullseye’). We found that total foliar flavonoids increased in response to UV due to flavonol upregulation, but only one class of flavonols, fisetin, exhibited stronger plasticity in alpine populations. Alpine populations tended to increase the quercetin-kaempferol ratio more than low elevation populations when exposed to higher UV, a signature of photoprotection and radical scavenging. Relationships between foliar flavonoids and the floral UV bullseye differed between alpine and low elevation populations. Previous work showed kaempferol glycosides contributed to variation in UV bullseye size at high elevation, while non-kaempferols spanning multiple FBP branches were associated with bullseye size at low elevation. Here, we found that alpine plants with less foliar kaempferol and greater kaempferol allocation to petals than leaves had larger floral UV-bullseyes, suggesting that floral UV patterning may be shaped by a biochemical tradeoff between tissues. Overall, nuanced elevational differences in flavonoid plasticity revealed by detailed metabolite classification provided support for local adaptation. Additionally, our study highlights that flavonoid production in leaves could influence the evolution of flavonoid-based floral phenotypes.

Proteomic analysis identified proteins that are differentially expressed in the flavonoid and carotenoid biosynthetic pathways of Camellia Nitidissima flowers

BackgroundCamellia nitidissima Chi is a popular ornamental plant because of its golden flowers, which contain flavonoids and carotenoids. To understand the regulatory mechanism of golden color formation, the metabolites of C. nitidissima petals at five different developmental stages were detected, a proteome map of petals was first constructed via tandem mass tag (TMT) analysis, and the accuracy of the sequencing data was validated via parallel reaction monitoring (PRM).ResultsNineteen color components were detected, and most of these components were carotenoids that gradually accumulated, while some metabolites were flavonoids that were gradually depleted. A total of 97,647 spectra were obtained, and 6,789 quantifiable proteins were identified. Then, 1,319 differentially expressed proteins (DEPs) were found, 55 of which belong to the flavonoid and carotenoid pathways, as revealed by pairwise comparisons of protein expression levels across the five developmental stages. Notably, most DEPs involved in the synthesis of flavonoids, such as phenylalanine ammonium lyase and 4-coumarate-CoA ligase, were downregulated during petal development, whereas DEPs involved in carotenoid synthesis, such as phytoene synthase, 1-deoxy-D-xylulose-5-phosphate synthase, and β-cyclase, tended to be upregulated. Furthermore, protein‒protein interaction (PPI) network analysis revealed that these 55 DEPs formed two distinct PPI networks closely tied to the flavonoid and carotenoid synthesis pathways. Phytoene synthase and chalcone synthase exhibited extensive interactions with numerous other proteins and displayed high connectivity within the PPI networks, suggesting their pivotal biological functions in flavonoid and carotenoid biosynthesis.ConclusionProteomic data on the flavonoid and carotenoid biosynthesis pathways were obtained, and the regulatory roles of the DEPs were analyzed, which provided a theoretical basis for further understanding the golden color formation mechanism of C. nitidissima.

Metabolome and transcriptome reveal the biosynthesis of flavonoids and amino acids in Isatis indigotica fruit during development.

Isatis indigotica Fort. is a famous medicinal plant that is also used as a natural dye and functional vegetable. The characteristics of the I. indigotica fruit during development are largely unknown, information that is essential for the exploitation and seedlings cultivation of I. indigotica. In this study, the biochemical, metabolite characteristics and gene expression profiling of I. indigotica at four developmental stages were investigated. A total of 428 metabolites were detected and categorized into 17 categories. High contents of anthocyanins, especially cyanidin 3-glucoside, might contribute to the purple colouration of I. indigotica fruits. Moreover, dozens of flavonoid components, including taxifolin, quercetin, astragalin and isovitexin 2″-O-beta-D-glucoside, and several other active components were also up-regulated in mature fruits. The abundance of antioxidants might endow a significantly stronger antioxidant activity of mature I. indigotica fruits compared to many other reported species. Enrichment analyses revealed that flavonoid and anthocyanin biosynthesis genes were mostly enriched in up-regulated gene sets during fruit development. The up-regulated structural genes, including IiCHS, IiCHI, IiF3H, IiDFR, IiANS, IiFLS, IiUGT, and transcription factors such as IiMYBs, IibHLHs and IiNACs were identified as candidate regulators of flavonoid and anthocyanin biosynthetic pathway. Furthermore, biosynthesis of amino acids was enriched in all pairwise comparisons of metabolites in fruits at four developmental stages. The differential accumulation of amino acids might result from the differentially expressed genes involved in amino acid biosynthesis. Taken together, these findings provide a comprehensive understanding of metabolite profiling and gene expression patterns in I. indigotica fruit during maturity, which is useful for pharmaceutical extractions and seedling cultivation of I. indigotica.

Identification of novel inhibitors of plant GH3 IAA-amido synthetases through molecular docking studies.

Auxins play a critical role in several plant developmental processes and their endogenous levels are regulated at multiple levels. The enzymes of the GRETCHEN HAGEN 3 (GH3) protein family catalyze the conjugation of amino acids to indoleacetic acid (IAA), the major endogenous auxin. The GH3 proteins are encoded by multiple redundant genes in plant genomes, making it difficult to perform functional genetic studies to understand their role in auxin homeostasis. To address these challenges, we used a chemical approach that exploits the reaction mechanism of GH3 proteins to identify small molecule inhibitors of their activity from a defined chemical library. The study evaluated receptor-ligand complexes based on their binding energy and classified them accordingly. Docking algorithms were used to correct any deviations, resulting in a list of the most important inhibitory compounds for selected GH3 enzymes based on a normalized sum of energy. The study presents atomic details of protein-ligand interactions and quantifies the effect of several of the identified small molecule inhibitors on auxin-mediated root growth processes in Arabidopsis thaliana. The direct effect of these compounds on endogenous auxin levels was measured using appropriate auxin sensors and endogenous hormone measurements. Our study has identified novel compounds of the flavonoid biosynthetic pathway that are effective inhibitors of GH3 enzyme-mediated IAA conjugation. These compounds play a versatile role in hormone-regulated plant development and have potential applications in both basic research and agriculture.

Chemical composition determination and transcriptomic analyses provide insight into the differences between wild and grafted Semen Ziziphi Spinosae

Semen Ziziphi Spinosae (SZS) is a traditional Chinese herbal medicine widely used to treat insomnia and anxiety in clinical practice. Currently, the demand for SZS is increasing every year, but the production of wild SZS is unstable due to environmental factors. Grafting sour jujube scions onto sour jujube or jujube tree stocks can achieve a high production rate within a short period of time. However, the effects of grafting on the quality of SZS have not been reported. This study investigated the differences between wild-type and grafted SZS from three aspects: phenotype, chemical composition, and molecular mechanism. The findings revealed that the grafted specimens were generally larger in morphology and lighter in color than the wild-type samples. The dimensions of both the grafted specimens were generally larger than those of the wild specimens. The HPLC-ELSD results revealed that the three main chemical components in the grafted SZS, namely, spinosin, jujuboside A, and jujuboside B, had higher contents than their wild-type counterparts. Comprehensive transcriptome sequencing analysis and KEGG annotation revealed that DEG enrichment between grafted and wild-type SZS occurred mainly during stress resistance and rootstock scion healing. There were 23 DEGs that may encode enzymes involved in the biosynthetic pathway of flavonoids and 21 genes encoding terpenoid saponins. Further investigation revealed that the expression of the genes C4H, CHS, CHI, and F3’5’H in the flavonoid biosynthesis pat.hway and HMGR, MVK, MVD, and FPPS in the saponin biosynthesis pathway accounted for the difference in quality between grafted and wild SZS. Furthermore, WGCNA identified 15 core genes related to medicinal ingredients between grafted and wild SZS. These results provide support for further research on the differences in the quality of medicinal ingredients between grafted and wild SZS.

Integrated Metabolome and Transcriptome Analyses Reveal the Mechanisms Regulating Flavonoid Biosynthesis in Blueberry Leaves under Salt Stress

The flavonoids play important roles in plant salt tolerance. Blueberries (Vaccinium spp.) are extremely sensitive to soil salt increases. Therefore, improving the salt resistance of blueberries by increasing the flavonoid content is crucial for the development of the blueberry industry. To explore the underlying molecular mechanism, we performed an integrated analysis of the metabolome and transcriptome of blueberry leaves under salt stress. We identified 525 differentially accumulated metabolites (DAMs) under salt stress vs. control treatment, primarily including members of the flavonoid class. We also identified 20,920 differentially expressed genes (DEGs) based on transcriptome data; of these, 568 differentially expressed transcription factors (TFs) were annotated, and bHLH123, OsHSP20, and HSP20 TFs might be responsible for blueberry leaf salt tolerance. DEGs involved in the flavonoid biosynthesis pathway were significantly enriched at almost all stages of salt stress. Salt treatment upregulated the expression of most flavonoid biosynthetic pathway genes and promoted the accumulation of flavonols, flavonol glycosides, flavans, proanthocyanidins, and anthocyanins. Correlation analysis suggested that 4-coumarate CoA ligases (4CL5 and 4CL1) play important roles in the accumulation of flavonols (quercetin and pinoquercetin) and flavan-3-ol (epicatechin and prodelphinidin C2) under salt stress, respectively. The flavonoid 3′5′-hydroxylases (F3′5′H) regulate anthocyanin (cyanidin 3-O-beta-D-sambubioside and delphinidin-3-O-glucoside chloride) biosynthesis, and leucoanthocyanidin reductases (LAR) are crucial for the biosynthesis of epicatechin and prodelphinidin C2 during salt stress. Taken together, it is one of the future breeding goals to cultivate salt-resistant blueberry varieties by increasing the expression of flavonoid biosynthetic genes, especially 4CL, F3′5′H, and LAR genes, to promote flavonoid content in blueberry leaves.

Identification of Black Cumin (Nigella sativa) MicroRNAs by Next-Generation Sequencing and Their Implications in Secondary Metabolite Biosynthesis.

Secondary metabolites are bioactive compounds believed to contribute to the pharmacological properties of plants. MicroRNAs (miRNAs) are small non-coding RNA molecules involved in post-transcriptional regulation and are thought to play an important role in regulating secondary metabolism biosynthesis. Nevertheless, the extent of miRNA involvement in secondary metabolism remains minimal. Nigella sativa (black cumin/black seed) is a popular medicinal and culinary plant known for its pharmaceutical properties; however, its genomic information is scarce. In this study, next-generation sequencing (NGS) technology was employed to obtain the miRNA profile of N. sativa, and their involvement in secondary metabolite biosynthesis was explored. A total of 25,139,003 unique reads ranging from 16 to 40 nucleotides were attained, out of which 240 conserved and 34 novel miRNAs were identified. Moreover, 6083 potential target genes were recognized in this study. Several conserved and novel black cumin miRNAs were found to target enzymes involved in the terpenoid, diterpenoid, phenylpropanoid, carotenoid, flavonoid, steroid, and ubiquinone biosynthetic pathways, among others, for example, beta-carotene 3-hydroxylase, gibberellin 3 beta-dioxygenase, trimethyltridecatetraene synthase, carboxylic ester hydrolases, acetyl-CoA C-acetyltransferase, isoprene synthase, peroxidase, shikimate O-hydroxycinnamoyltransferase, etc. Furthermore, sequencing data were validated through qPCR by checking the relative expression of eleven randomly selected conserved and novel miRNAs (nsa-miR164d, nsa-miR166a, nsa-miR167b, nsa-miR171a, nsa-miR390b, nsa-miR396, nsa-miR159a, nsa-miRN1, nsa-miRN29, nsa-miRN32, and nsa-miRN34) and their expression patterns were found to be corroborated with the sequencing data. We anticipate that this work will assist in clarifying the implications of miRNAs in plant secondary metabolism and aid in the generation of artificial miRNA-based strategies to overproduce highly valuable secondary metabolites from N. sativa.

Chromosome-scale genome assembly of Astragalus membranaceus using PacBio and Hi-C technologies

Astragalus membranaceus (Fisch.) Bge (AM) is a medicinal herb plant belonging to the Leguminosae family. In this study, we present a chromosome-scale genome assembly of AM, aiming to enhance the molecular biology and functional studies of Astragali Radix. The genome size of AM is about 1.43 Gb, with a contig N50 value of 1.67 Mb. A total of 98.16% of the assembly anchored to 9 pseudochromosomes using Hi-C technology. The assembly completeness was estimated to be 97.27% using BUSCO with the long terminal repeat assembly index (LAI) of 16.22 and quality value (QV) of 48.58. Additionally, the genome contained 67.98% repetitive sequences. Genome annotation predicted 29,914 protein-coding genes, including 73 genes involved in the flavonoid biosynthetic pathway and 2,048 transcription factors. The high-quality genome assembly and gene annotation resources will greatly facilitate future functional genomic studies in Leguminosae species.

FtMYB163 Gene Encodes SG7 R2R3-MYB Transcription Factor from Tartary Buckwheat (Fagopyrum tataricum Gaertn.) to Promote Flavonol Accumulation in Transgenic Arabidopsis thaliana.

Tartary buckwheat (Fagopyrum tataricum Gaertn.) is a coarse grain crop rich in flavonoids that are beneficial to human health because they function as anti-inflammatories and provide protection against cardiovascular disease and diabetes. Flavonoid biosynthesis is a complex process, and relatively little is known about the regulatory pathways involved in Tartary buckwheat. Here, we cloned and characterized the FtMYB163 gene from Tartary buckwheat, which encodes a member of the R2R3-MYB transcription factor family. Amino acid sequence and phylogenetic analysis indicate that FtMYB163 is a member of subgroup 7 (SG7) and closely related to FeMYBF1, which regulates flavonol synthesis in common buckwheat (F. esculentum). We demonstrated that FtMYB163 localizes to the nucleus and has transcriptional activity. Expression levels of FtMYB163 in the roots, stems, leaves, flowers, and seeds of F. tataricum were positively correlated with the total flavonoid contents of these tissues. Overexpression of FtMYB163 in transgenic Arabidopsis enhanced the expression of several genes involved in early flavonoid biosynthesis (AtCHS, AtCHI, AtF3H, and AtFLS) and significantly increased the accumulation of several flavonoids, including naringenin chalcone, naringenin-7-O-glucoside, eriodictyol, and eight flavonol compounds. Our findings demonstrate that FtMYB163 positively regulates flavonol biosynthesis by changing the expression of several key genes in flavonoid biosynthetic pathways.

Transcriptomic and coexpression network analyses revealed the regulatory mechanism of Cydia pomonella infestation on the synthesis of phytohormones in walnut husks

The codling moth (Cydia pomonella) has a major effect on the quality and yield of walnut fruit. Plant defences respond to insect infestation by activating hormonal signalling and the flavonoid biosynthetic pathway. However, little is known about the role of walnut husk hormones and flavonoid biosynthesis in response to C. pomonella infestation. The phytohormone content assay revealed that the contents of salicylic acid (SA), abscisic acid (ABA), jasmonic acid (JA), jasmonic acid-isoleucine conjugate (JA-ILE), jasmonic acid-valine (JA-Val) and methyl jasmonate (MeJA) increased after feeding at different time points (0, 12, 24, 36, 48, and 72 h) of walnut husk. RNA-seq analysis of walnut husks following C. pomonella feeding revealed a temporal pattern in differentially expressed genes (DEGs), with the number increasing from 3,988 at 12 h to 5,929 at 72 h postfeeding compared with the control at 0 h postfeeding. Walnut husks exhibited significant upregulation of genes involved in various defence pathways, including flavonoid biosynthesis (PAL, CYP73A, 4CL, CHS, CHI, F3H, ANS, and LAR), SA (PAL), ABA (ZEP and ABA2), and JA (AOS, AOC, OPR, JAZ, and MYC2) pathways. Three gene coexpression networks that had a significant positive association with these hormonal changes were constructed based on the basis of weighted gene coexpression network analysis (WGCNA). We identified several hub transcription factors, including the turquoise module (AIL6, MYB4, PRE6, WRKY71, WRKY31, ERF003, and WRKY75), the green module (bHLH79, PCL1, APRR5, ABI5, and ILR3), and the magenta module (ERF27, bHLH35, bHLH18, TIFY5A, WRKY31, and MYB44). Taken together, these findings provide useful genetic resources for exploring the defence response mediated by phytohormones in walnut husks.

McWRKY43 Confers Cold Stress Tolerance in Michelia crassipes via Regulation of Flavonoid Biosynthesis.

WRKY transcription factor (TF) plays a crucial role in plant abiotic stress response, but it is rarely reported in Michelia crassipes. Our studies have found that the transcription factor McWRKY43, a member of the IIc subgroup, is strongly upregulated under cold stress. In this study, we cloned the full length of McWRKY43 to further investigate the function of McWRKY43 in resistance to cold stress and its possible regulatory pathways in M. crassipes. Under cold stress, the seed-germination rate of transgenic tobacco was significantly higher than that of the wild type, and the flavonoid content, antioxidant enzyme activities, and proline content of transgenic tobacco seedlings were significantly increased, which promoted the expression of flavonoid pathway structural genes. In addition, the transient transformation of McWRKY43 in the M. crassipes leaves also found the accumulation of flavonoid content and the transcription level of flavonoid structural genes, especially McLDOX, were significantly increased under cold stress. Yeast one-hybrid (Y1H) assay showed that McWRKY43 could bind to McLDOX promoter, and the transcription expression of McLDOX was promoted by McWRKY43 during cold stress treatment. Overall, our results indicated that McWRKY43 is involved in flavonoid biosynthetic pathway to regulate cold stress tolerance of M. crassipes, providing a basis for molecular mechanism of stress resistance in Michelia.

Genome-wide analysis of flavonoid biosynthetic genes in Musaceae (Ensete, Musella, and Musa species) reveals amplification of flavonoid 3ʹ,5ʹ-hydroxylase

Abstract Flavonoids in Musaceae are involved in pigmentation and stress responses, including cold resistance, and are a component of the healthy human diet. Identification and analysis of the sequence and copy number of flavonoid biosynthetic genes are valuable for understanding the nature and diversity of flavonoid evolution in Musaceae species. In this study, we identified 71–80 flavonoid biosynthetic genes in chromosome-scale genome sequence assemblies of Musaceae, including those of Ensete glaucum, Musella lasiocarpa, Musa beccarii, M. acuminata, M. balbisiana and M. schizocarpa, checking annotations with BLAST and determining the presence of conserved domains. The number of genes increased through segmental duplication and tandem duplication. Orthologues of both structural and regulatory genes in the flavonoid biosynthetic pathway are highly conserved across Musaceae. The flavonoid 3ʹ,5ʹ-hydroxylase gene F3ʹ5ʹH was amplified in Musaceae and ginger compared with grasses (rice, Brachypodium, Avena longiglumis, and sorghum). One group of genes from this gene family amplified near the centromere of chromosome 2 in the x = 11 Musaceae species. Flavonoid biosynthetic genes displayed few consistent responses in the yellow and red bracts of Musella lasiocarpa when subjected to low temperatures. The expression levels of MlDFR2/3 (dihydroflavonol reductase) increased while MlLAR (leucoanthocyanidin reductase) was reduced by half. Overall, the results establish the range of diversity in both sequence and copy number of flavonoid biosynthetic genes during evolution of Musaceae. The combination of allelic variants of genes, changes in their copy numbers, and variation in transcription factors with the modulation of expression under cold treatments and between genotypes with contrasting bract-colours suggests the variation may be exploited in plant breeding programmes, particularly for improvement of stress-resistance in the banana crop.

Comprehensive genomic analysis and expression profiling of the cytochrome P450 genes during abiotic stress and flavonoid biosynthesis in potato (Solanum tuberosum)

The research focused on the cytochrome P450 (CYP) superfamily, a prominent enzyme family involved in plant metabolism. A total of 253 StCYP genes from the potato genome were investigated and characterized through various methods, including phylogentic analysis, physical and chemical characterization, chromosome localization, gene structure and conserved protein domain classification, gene duplication occurrences, collinearity and expression pattern analysis. The 253 StCYPs were classified into seven clans and 33 families, distributed unevenly across 12 chromosomes. Collinearity analysis revealed 28 pairs of orthologous genes between potato StCYPs and Arabidopsis AtCYPs. Moreover, the expression of the StCYPs in various tissue parts of doubled haploid (DM) potatoes under abiotic stress and exogenous hormone treatment was investigated by using RNA-seq data from the PGSC database. It was discovered that one StCYP gene likely contributes to drought stress response through the ABA-dependent pathway. Furthermore, RNA-seq analysis of pigmented tuber flesh from three potato clones at three developmental stages enabled the identification of StCYPs potentially implicated in the flavonoid biosynthesis in potato tubers. In particular, StCYP705A-like was further validated for its potential role in this process. Employing Gfanno software in conjunction with RNA-seq data, seven StCYP genes (Two StC4H, three StF3′H, one StFNS Ⅱ and one StF3′5′H) were identified as flavonoid biosynthetic pathway genes in potato. Additionally, five StCYPs in CYP 71 clan demonstrated a strong association with flavonoid biosynthesis. These findings lay a groundwork for a more comprehensive understanding of the StCYP gene family and further exploration of the functions of StCYP genes in potato abiotic stress tolerance and flavonoids biosynthesis.

Regulation of exogenous sugars on the biosynthesis of key secondary metabolites in Cyclocarya paliurus.

The biosynthesis and accumulation of secondary metabolites play a vital role in determining the quality of medicinal plants, with carbohydrate metabolism often influencing secondary metabolism. To understand the potential regulatory mechanism, exogenous sugars (sucrose, glucose/fructose) were applied to the leaves of Cyclocarya paliurus, a highly valued and multiple function tree species. The results showed that exogenous sugars enhanced the accumulation of soluble sugar and starch while increasing the enzyme activity related to carbohydrate metabolism. In addition, the plant height was increased by a mixture of exogenous mixed sugars, the addition of sucrose promoted the net photosynthetic rate, while all types of exogenous sugars facilitated the accumulation of flavonoids and terpenoids. Based on weighted gene co-expression network analysis (WGCNA), two key gene modules and four candidate transcription factors (TFs) related to carbohydrate metabolism and secondary metabolite biosynthesis were identified. A correlation analysis between transcriptome and metabolome data showed that exogenous sugar up-regulated the expression of key structural genes in the flavonoid and terpenoid biosynthetic pathway. The expression levels of the four candidate TFs, TIFY 10A, WRKY 7, EIL 3 and RF2a, were induced by exogenous sugar and were strongly correlated with the key structural genes, which enhanced the synthesis of specific secondary metabolites and some plant hormone signal pathways. Our results provide a comprehensive understanding of key factors in the quality formation of medicinal plants and a potential approach to improve the quality.

Metabolomic analysis combined with machine learning algorithms enables the evaluation of postharvest pecan color stability

Nut kernel color is a crucial quality indicator affecting the consumers first impression of the product. While growing evidence suggests that plant phenolics and their derivatives are linked to nut kernel color, the compounds (biomarkers) responsible for kernel color stability during storage remain elusive. Here, pathway-based metabolomics with machine learning algorithms were employed to identify key metabolites of postharvest pecan color stability. Metabolites in phenylpropanoid, flavonoid, and anthocyanin biosynthetic pathways were analyzed in the testa of nine pecan cultivars using liquid chromatography–mass spectrometry. With color measurements, different machine learning models were compared to find relevant biomarkers of pecan color phenotypes. Results revealed potential marker compounds that included flavonoid precursors and anthocyanidins as well as anthocyanins (e.g., peonidin, delphinidin-3-O-glucoside). Our findings provide a foundation for future research in the area, and will help select genes/proteins for the breeding of pecans with stable and desirable kernel color.

Identification of key genes involved in lignin and flavonoid accumulation during Tilia tuan seed maturation.

Transcriptomics and phenotypic data analysis identified 24 transcription factors (TFs) that play key roles in regulating the competitive accumulation of lignin and flavonoids. Tilia tuan Szyszyl. (T. tuan) is a timber tree species with important ecological and commercial value. However, its highly lignified pericarp results in a low seed germination rate and a long dormancy period. In addition, it is unknown whether there is an interaction between the biosynthesis of flavonoids and lignin as products of the phenylpropanoid pathway during seed development. To explore the molecular regulatory mechanism of lignin and flavonoid biosynthesis, T. tuan seeds were harvested at five stages (30, 60, 90, 120, and 150 days after pollination) for lignin and flavonoid analyses. The results showed that lignin accumulated rapidly in the early and middle stages (S1, S3, and S4), and rapid accumulation of flavonoids during the early and late stages (S1 and S5). High-throughput RNA sequencing analysis of developing seeds identified 50,553 transcripts, including 223 phenylpropanoid biosynthetic pathway genes involved in lignin accumulation grouped into 3 clusters, and 106 flavonoid biosynthetic pathway genes (FBPGs) grouped into 2 clusters. Subsequent WGCNA and time-ordered gene co-expression network (TO-GCN) analysis revealed that 24 TFs (e.g., TtARF2 and TtWRKY15) were involved in flavonoids and lignin biosynthesis regulation. The transcriptome data were validated by qRT-PCR to analyze the expression profiles of key enzyme-coding genes. This study revealed that there existed a competitive relationship between flavonoid and lignin biosynthesis pathway during the development of T. tuan seeds, that provide a foundation for the further exploration of molecular mechanisms underlying lignin and flavonoid accumulation in T. tuan seeds.

Citric acid treatment inhibits fading of sorghum (Sorghum bicolor) by modulating the accumulation of flavonoids

Sorghum seeds can discolor during storage. Treatment of seeds with citric acid improves sensory quality and antioxidant activity. This study compared the differences in phenotypic and antioxidant activity between citric acid-treated and water-treated sorghum seeds. The study used transcriptomics and metabolomics approaches to investigate the regulatory mechanisms. The ∆a, ∆b and ∆l values of citric acid-treated sorghum seeds significantly increased after 6 months of storage. The SOD, POD and CAT enzyme activities of the citric acid-treated group were 1.94, 1.91 and 2.45 times higher than those of the control, respectively. The joint transcriptome and metabolome analysis showed that the citric acid-induced changes were mainly focused on the flavonoid biosynthetic pathway. Citric acid treatment up-regulated CHS, ANR, MYB and bHLH genes and promoted flavonoid accumulation. In conclusion, citric acid treatment promotes flavonoid accumulation, delays sorghum seed discoloration, and enhances antioxidant activity and storage life.

The gap-free genome and multi-omics analysis of Citrus reticulata 'Chachi' reveal the dynamics of fruit flavonoid biosynthesis.

Citrus reticulata 'Chachi' (CRC) has long been recognized for its nutritional benefits, health-promoting properties, and pharmacological potential. Despite its importance, the bioactive components of CRC and their biosynthetic pathways have remained largely unexplored. In this study, we introduce a gap-free genome assembly for CRC, which has a size of 312.97Mb and a contig N50 size of 32.18Mb. We identified key structural genes, transcription factors, and metabolites crucial to flavonoid biosynthesis through genomic, transcriptomic, and metabolomic analyses. Our analyses reveal that 409 flavonoid metabolites, accounting for 83.30% of the total identified, are highly concentrated in the early stage of fruit development. This concentration decreases as the fruit develops, with a notable decline in compounds such as hesperetin, naringin, and most polymethoxyflavones observed in later fruit development stages. Additionally, we have examined the expression of 21 structural genes within the flavonoid biosynthetic pathway, and found a significant reduction in the expression levels of key genes including 4CL, CHS, CHI, FLS, F3H, and 4'OMT during fruit development, aligning with the trend of flavonoid metabolite accumulation. In conclusion, this study offers deep insights into the genomic evolution, biosynthesis processes, and the nutritional and medicinal properties of CRC, which lay a solid foundation for further gene function studies and germplasm improvement in citrus.