Search Results (51)

Flavonoids serve as crucial plant antioxidants in drought tolerance, yet their antioxidant regulatory mechanisms within mycorrhizal plants remain unclear. In this study, using a two-factor design, trifoliate orange (Poncirus trifoliata (L.) Raf.) seedlings in the four-to-five-leaf stage were either inoculated with Funneliformis mosseae or not, and subjected to well-watered (70–75% of field maximum water-holding capacity) or drought stress (50–55% field maximum water-holding capacity) conditions for 10 weeks. Plant growth performance, photosynthetic physiology, leaf flavonoid content and their antioxidant capacity, reactive oxygen species levels, and activities and gene expression of key flavonoid biosynthesis enzymes were analyzed. Although drought stress significantly reduced root colonization and soil hyphal length, inoculation with F. mosseae consistently enhanced the biomass of leaves, stems, and roots, as well as root surface area and diameter, irrespective of soil moisture. Despite drought suppressing photosynthesis in mycorrhizal plants, F. mosseae substantially improved photosynthetic capacity (measured via gas exchange) and optimized photochemical efficiency (assessed by chlorophyll fluorescence) while reducing non-photochemical quenching (heat dissipation). Inoculation with F. mosseae elevated the total flavonoid content in leaves by 46.67% (well-watered) and 14.04% (drought), accompanied by significantly enhanced activities of key synthases such as phenylalanine ammonia-lyase (PAL), chalcone synthase (CHS), chalcone isomerase (CHI), 4-coumarate:coA ligase (4CL), and cinnamate 4-hydroxylase (C4H), with increases ranging from 16.90 to 117.42% under drought. Quantitative real-time PCR revealed that both mycorrhization and drought upregulated the expression of PtPAL1, PtCHI, and Pt4CL genes, with soil moisture critically modulating mycorrhizal regulatory effects. In vitro assays showed that flavonoid extracts scavenged radicals at rates of 30.07–41.60% in hydroxyl radical (•OH), 71.89–78.06% in superoxide radical anion (O₂^•−), and 49.97–74.75% in 2,2-diphenyl-1-picrylhydrazyl (DPPH). Mycorrhizal symbiosis enhanced the antioxidant capacity of flavonoids, resulting in higher scavenging rates of •OH (19.07%), O₂^•− (5.00%), and DPPH (31.81%) under drought. Inoculated plants displayed reduced hydrogen peroxide (19.77%), O₂^•− (23.90%), and malondialdehyde (17.36%) levels. This study concludes that mycorrhizae promote the level of total flavonoids in trifoliate orange by accelerating the flavonoid biosynthesis pathway, hence reducing oxidative damage under drought. Full article

The occurrence of anthocyanins in rice (Oryza sativa) and barley (Hordeum vulgare) varies among cultivars, with pigmented varieties (e.g., black rice and purple barley) accumulating higher concentrations due to genetic and environmental factors. The biosynthesis of anthocyanins is regulated by a complex network of structural and regulatory genes. Key enzymes in the pathway include chalcone synthase (CHS), chalcone isomerase (CHI), flavanone 3-hydroxylase (F3H), dihydroflavonol 4-reductase (DFR), anthocyanidin synthase (ANS), and UDP-glucose flavonoid 3-O-glucosyltransferase (UFGT). These genes are tightly controlled by transcription factors (TFs) from the MYB, bHLH (basic helix–loop–helix), and WD40 repeat families, which form the MBW (MYB-bHLH-WD40) regulatory complex. In rice, OsMYB transcription factors such as OsMYB3, OsC1, and OsPL (Purple Leaf) interact with OsbHLH partners (e.g., OsB1, OsB2) to activate anthocyanin biosynthesis. Similarly, in barley, HvMYB genes (e.g., HvMYB10) coordinate with HvbHLH TFs to regulate pigment accumulation. Environmental cues, such as light, temperature, and nutrient availability, further modulate these TFs, influencing the production of anthocyanin. Understanding the genetic and molecular mechanisms behind the biosynthesis of anthocyanins in rice and barley provides opportunities for the development of biofortification strategies that enhance their nutritional value. Full article

Flavonoid synthases (FNSs) are key enzymes catalyzing the conversion of flavanones to flavonoids, yet their functions in citrus remain functionally uncharacterized. In this study, we identified three FNSII genes in the citrus genome. Phylogenetic analysis revealed that citrus FNSII genes share the closest evolutionary distance with apple FNSII genes. Chromosomal localization demonstrated that the three FNSII genes are distributed across two out of nine chromosomes. Gene structure analysis indicated that the majority of motifs within these three FNSII genes are highly conserved. We cloned a gene called CitFNSII-1 from citrus. Transient overexpression of CitFNSII-1 in citrus leaves significantly increased flavonoid content, while simultaneous virus-induced silencing of CitFNSII-1 led to synchronously and significantly reduced gene expression levels and flavonoid content in citrus seedlings. Through the Agrobacterium rhizogenes-mediated genetic transformation system, overexpression of CitFNSII-1 was found to markedly enhance flavonoid accumulation in hairy roots, whereas knockout of CitFNSII-1 resulted in a significant decrease in flavonoid content in hairy roots. Further experiments verified an interaction between CitFNSII-1 and the Chalcone isomerase-1 (CHI-1) protein. The results demonstrated that the flavonoid accumulation patterns of CHI-1 and CitFNSII-1 are highly similar. In conclusion, this study advances the understanding of the flavonoid biosynthesis pathway in citrus and provides a theoretical foundation for molecular breeding strategies in citrus. Full article

The seed-coat color and seed size have an impact on both the evolutionary fitness and the grain yield of crops. Soybean is a major oil crop, and the seed-coat color and seed size exhibit natural diversity among the different soybean varieties. Here, we found an R2R3-MYB transcription factor of GmMYB62, which shows a significant increase in expression as the seed-coat color changes from yellow to black in different soybean varieties. The GmMYB62 was specifically highly expressed in reproductive organs, especially in floral organs in soybeans. The GmMYB62 encodes a nuclear protein that contains two MYB domains. In the phylogenetic analysis, the GmMYB62 was relatively conserved after the divergence of the monocots and dicots, and it also grouped with transcriptional repressors of MYBs in anthocyanin synthesis. The GmMYB62 was overexpressed in Arabidopsis and the seeds displayed a pale-brown coat in GmMYB62 overexpression lines, in contrast to the dark-brown seed coat observed in wild-type of Col-0. The anthocyanin content in the GmMYB62 overexpression lines was dramatically reduced when compared to Col-0. Additionally, the seeds in overexpression lines showed shorter lengths, larger widths, and lower thousand-seed weights than those in Col-0. Furthermore, the genes related to anthocyanin synthesis and seed size regulation were investigated, and expression of eight genes that involved in anthocyanin synthesis pathway, like chalcone synthase (CHS), chalcone isomerase (CHI), flavanone 3-hydroxylase (F3H), and anthocyanidin synthase (ANS) were severely inhibited in the GmMYB62 overexpression lines when compared to Col-0. In addition, the ARGOS-LIKE (ARL), B-Type Cyclin 1 (CYCB1), and enhancer of DA1-1 (EOD3), which govern cell expansion and proliferation, were highly expressed in GmMYB62 overexpression lines when compared to Col-0. Overall, this study sheds new light on the control of seed-coat color and seed size by GmMYB62 and provides potentially valuable targets for improving crop seed quality. Full article

Chalcone isomerase (CHI) is not only an enzyme related to flavonoid biosynthesis, but also one of the key enzymes in the flavonoid metabolic pathway. In this study, members of the CHI gene family were identified in the whole genome of sweet potato. Bioinformatics methods were used to analyze the physical and chemical properties, systematic evolution, conserved domain, chromosome location, cis-acting elements of the promoter, and so on, of CHI gene family members. In addition, the tissue site-specific expression of CHI gene family members and their expression patterns under three kinds of abiotic stress were analyzed. The results showed that five members of IbCHI gene family were identified in sweet potato, which were unevenly distributed on four chromosomes. The protein secondary structure and tertiary structure were consistent, and there was a conservative domain related to chalcone isomerase. The prediction of subcellular localization showed that it was mainly located in cytoplasm and chloroplast. Systematic evolution showed that the members of sweet potato CHI gene family could be divided into Type I-IV, and the Type I gene IbCHI1 showed CHI catalytic activity in transgenic callus. The collinearity gene pairs were identified between sweet potato and allied species. Its promoter contains light response elements, hormone response elements, and stress response elements. The results of real-time fluorescence quantitative PCR (qRT-PCR) analysis showed that the expression of the IbCHI gene was tissue-specific and that the catalytic genes IbCHI1 and IbCHI5 serve as primary responders to abiotic stress, while the non-catalytic members IbCHI3 and IbCHI4 may fine-tune metabolic flux or participate in low-temperature, salt, and drought stress signaling. This study can provide a theoretical basis for a follow-up functional genomics study of the chalcone isomerase gene family in sweet potato. Full article

Waxy maize (Zea mays L. sinensis kulesh) contains a lot of nutrients, and has a long history of cultivation and extensive consumption. In this study, six waxy maize varieties of white (J18 and W2000), yellow (J41 and J7), and black (J10 and J20) were selected as experimental materials, and the functional nutrients and the differences in anthocyanin anabolic pathways in maize kernels at 14, 18, 22, and 26 days after pollination were determined. The result show that the varieties and kernel development stages had significant effect on the carotenoid, soluble sugar, vitamin C, anthocyanin, and mineral element content. The black waxy maize varieties had a higher anthocyanin content, which plays an important role in maize kernel coloration, whereas the yellow and black waxy maize varieties exhibit a greater abundance of mineral elements. Furthermore, the phenylalanine content, as well as the activities of phenylalanine ammonia lyase (PAL), chalcone isomerase (CHI), dihydroflavonol reductase (DFR), and flavonoid 3-glucosyltransferase (UFGT), played a significant role in the anthocyanin biosynthetic pathway. In conclusion, the comprehensive functional quality of waxy maize decreased with the delay of kernel development stage, and the black waxy maize varieties demonstrated superior functional quality. The PAL and CHI played a primary role in the initial phase of anthocyanin accumulation, while UFGT gradually assumed control in the subsequent stages. Full article

White clover (Trifolium repens) is vulnerable to drought stress. In response to abiotic stress, plants are regulated by NAC transcription factors. The NAC in white clover has not been thoroughly documented until recently. We have identified one white clover NAC transcription factor called TrNAC002. TrNAC002’s coding sequence is localized to specific regions on the 3P and 5O chromosomes of white clover and is part of a single-copy nuclear gene. Subcellular localization demonstrates that TrNAC002 is located in the nucleus, while the transcriptional activity assay indicates its transcriptional activity. Arabidopsis plants overexpressing TrNAC002 (OE) exhibit enlarged leaves and increased lateral root growth compared to the wild type (WT). Additionally, the expression levels of the shoot apical meristem (SAM), WUSCHEL (WUS), DNA-binding protein (DBP), and auxin-induced in root cultures3 (AIR3) genes are significantly higher in OE as compared to WT. These findings imply that TrNAC002 could promote vegetative growth by increasing the expression of these genes. Under natural drought stress, OE can survive in dry soil for a longer period of time than WT. Furthermore, OE exhibits a lower level of reactive oxygen species (ROS) level and a higher content of flavonoids than WT. This is also positively correlated with an increased flavonoid content. In white clover, the expression of TrNAC002, chalcone synthase (CHS), and chalcone isomerase (CHI) in leaves demonstrates significant upregulation after drought stress and ABA treatment, as does the flavonoid content. However, the pTRV-VIGS experiment suggests that pTRV2-TrNAC002 white clover shrinks compared to the Mock and Water controls. Additionally, pTRV2-TrNAC002 white clover displays a statistically higher malondialdehyde (MDA) content than the Mock and Water controls, and a significantly lower level of total antioxidant activities, flavonoid content, CHS and CHI relative expression than that of the Mock and Water controls. These findings indicate that TrNAC002 responds to drought and modulates flavonoid biosynthesis in white clover. This study is the first to suggest that TrNAC002 likely responds to drought via ABA and enhances plant drought resistance by synthesizing flavonoids. Full article

Elevated levels of reactive oxygen species (ROS) are caused by ultraviolet B radiation (UV-B) stress. In response, plants strengthen their cell membranes, impeding photosynthesis. Additionally, UV-B stress initiates oxidative stress within the antioxidant defense system and alters secondary metabolism, particularly by increasing the quantity of UV-absorbing compounds such as flavonoids. The v-myb avian myeloblastosis viral oncogene homolog (MYB) transcription factor (TF) may participate in a plant’s response to UV-B damage through its regulation of flavonoid biosynthesis. In this study, we discovered that the photosynthetic activity of Rhododendron chrysanthum Pall. (R. chrysanthum) decreased when assessing parameters of chlorophyll (PSII) fluorescence parameters under UV-B stress. Concurrently, antioxidant system enzyme expression increased under UV-B exposure. A multi-omics data analysis revealed that acetylation at the K68 site of the RcTRP5 (telomeric repeat binding protein of Rhododendron chrysanthum Pall.) transcription factor was upregulated. This acetylation modification of RcTRP5 activates the antioxidant enzyme system, leading to elevated expression levels of peroxidase (POD), superoxide dismutase (SOD), and catalase (CAT). Upregulation is also observed at the K95 site of the chalcone isomerase (CHI) enzyme and the K178 site of the anthocyanidin synthase (ANS) enzyme. We hypothesize that RcTRP5 influences acetylation modifications of CHI and ANS in flavonoid biosynthesis, thereby indirectly regulating flavonoid production. This study demonstrates that R. chrysanthum can be protected from UV-B stress by accumulating flavonoids. This could serve as a useful strategy for enhancing the plant’s flavonoid content and provide a valuable reference for research on the metabolic regulation mechanisms of other secondary substances. Full article

Soil salinization is a significant environmental stress factor, threatening global agricultural yield and ecological security. Plants must effectively cope with the adverse effects of salt stress on survival and successful reproduction. Lateral Organ Boundaries (LOB) Domain (LBD) genes, a gene family encoding plant-specific transcription factors (TFs), play important roles in plant growth and development. Here, we identified and functionally characterized the LBD family TF PtrLBD41 from Populus trichocarpa, which can be induced by various abiotic stresses, including salt, dehydration, low temperature, and Abscisic Acid (ABA). Meanwhile, transgenic plants overexpressing PtrLBD41 showed a better phenotype and higher tolerance than the wild-type (WT) plants under salt stress treatment. Transcriptome analysis found that the differentially expressed genes (DEGs) between the WT and overexpression (OE) line were enriched in the flavonoid biosynthetic process, in which chalcone synthases (CHS), naringenin 3-dioxygenase (F3H), and chalcone isomerase (CHI) were significantly up-regulated under salt stress conditions through qRT-PCR analysis. Therefore, we demonstrate that PtrLBD41 plays an important role in the tolerance to salt stress in P. trichocarpa. Full article

Camellia hainanica is one of the camellia plants distributed in tropical regions, and its regeneration system and genetic transformation are affected by callus browning. However, the underlying mechanism of Camellia hainanica callus browning formation remains largely unknown. To investigate the metabolic basis and molecular mechanism of the callus browning of Camellia hainanica, histological staining, high-throughput metabolomics, and transcriptomic assays were performed on calli with different browning degrees (T1, T2, and T3). The results of histological staining revealed that the brown callus cells had obvious lignification and accumulation of polyphenols. Widely targeted metabolomics revealed 1190 differentially accumulated metabolites (DAMs), with 53 DAMs annotated as phenylpropanoids and flavonoids. Comparative transcriptomics revealed differentially expressed genes (DEGs) of the T2 vs. T1 associated with the biosynthesis and regulation of flavonoids and transcription factors in Camellia hainanica. Among them, forty-four enzyme genes associated with flavonoid biosynthesis were identified, including phenylalaninase (PAL), 4-coumaroyl CoA ligase (4CL), naringenin via flavanone 3-hydroxylase (F3H), flavonol synthase (FLS), Chalcone synthase (CHS), Chalcone isomerase (CHI), hydroxycinnamoyl-CoA shikimate transferase (HCT), Dihydroflavonol reductase (DFR), anthocyanin reductase (LAR), anthocyanin synthetase (ANS), and anthocyanin reductase (ANR). Related transcription factors R2R3-MYB, basic helix-loop-helix (bHLH), and WRKY genes also presented different expression patterns in T2 vs. T1. These results indicate that the browning of calli in Camellia hainanica is regulated at both the transcriptional and metabolic levels. The oxidation of flavonoids and the regulation of related structural genes and transcription factors are crucial decisive factors. This study preliminarily revealed the molecular mechanism of the browning of the callus of Camellia hainanensis, and the results can provide a reference for the anti-browning culture of Camellia hainanica callus. Full article

Chrysanthemums are among the world’s most popular cut flowers, with their color being a key ornamental feature. The formation of these colors can be influenced by high temperatures. However, the regulatory mechanisms that control the fading of chrysanthemum flower color under high-temperature stress remain unclear. This study investigates the impact of high temperatures on the color and biochemical responses of purple chrysanthemums. Four purple chrysanthemum varieties were exposed to both normal and elevated temperature conditions. High-temperature stress elicited distinct responses among the purple chrysanthemum varieties. ‘Zi Feng Che’ and ‘Chrystal Regal’ maintained color stability, whereas ‘Zi Hong Tuo Gui’ and ‘Zi lian’ exhibited significant color fading, particularly during early bloom stages. This fading was associated with decreased enzymatic activities, specifically of chalcone isomerase (CHI), dihydroflavonol 4-reductase (DFR), and anthocyanidin synthase (ANS), indicating a critical period of color development under heat stress. Additionally, the color fading of ‘Zi Lian’ was closely related to the increased activity of the peroxidase (POD) and polyphenol oxidase (PPO). Conversely, a reduction in β-glucosidase (βG) activity may contribute significantly to the color steadfastness of ‘Zi Feng Che’. The genes Cse_sc027584.1_g010.1 (PPO) and Cse_sc031727.1_g010.1 (POD) might contribute to the degradation of anthocyanins in the petals of ‘Zi Hong Tuo Gui’ and ‘Zi Lian’ under high-temperature conditions, while simultaneously maintaining the stability of anthocyanins in ‘Zi Feng Che’ and ‘Chrystal Regal’ at the early bloom floral stage. The findings of this research provide new insights into the physiological and biochemical mechanisms by which chrysanthemum flower color responds to high-temperature stress. Full article

Chalcone synthase (CHS) and chalcone isomerase (CHI) catalyze the first two committed steps of the flavonoid pathway that plays a pivotal role in the growth and reproduction of land plants, including UV protection, pigmentation, symbiotic nitrogen fixation, and pathogen resistance. Based on the obtained X-ray crystal structures of CHS, CHI, and chalcone isomerase-like protein (CHIL) from the same monocotyledon, Panicum virgatum, along with the results of the steady-state kinetics, spectroscopic/thermodynamic analyses, intermolecular interactions, and their effect on each catalytic step are proposed. In addition, PvCHI’s unique activity for both naringenin chalcone and isoliquiritigenin was analyzed, and the observed hierarchical activity for those type-I and -II substrates was explained with the intrinsic characteristics of the enzyme and two substrates. The structure of PvCHS complexed with naringenin supports uncompetitive inhibition. PvCHS displays intrinsic catalytic promiscuity, evident from the formation of p-coumaroyltriacetic acid lactone (CTAL) in addition to naringenin chalcone. In the presence of PvCHIL, conversion of p-coumaroyl-CoA to naringenin through PvCHS and PvCHI displayed ~400-fold increased V_max with reduced formation of CTAL by 70%. Supporting this model, molecular docking, ITC (Isothermal Titration Calorimetry), and FRET (Fluorescence Resonance Energy Transfer) indicated that both PvCHI and PvCHIL interact with PvCHS in a non-competitive manner, indicating the plausible allosteric effect of naringenin on CHS. Significantly, the presence of naringenin increased the affinity between PvCHS and PvCHIL, whereas naringenin chalcone decreased the affinity, indicating a plausible feedback mechanism to minimize spontaneous incorrect stereoisomers. These are the first findings from a three-body system from the same species, indicating the importance of the macromolecular assembly of CHS-CHI-CHIL in determining the amount and type of flavonoids produced in plant cells. Full article

Chalcone isomerase (CHI) is an important enzyme involved in the biosynthesis of flavonoids, one that is crucial in both plant defense and human health. Although many CHI genes have been previously identified, the function of CHI-like genes in soybean remains unclear. In this study, we cloned the CHI-like genes GmCHI4A and GmCHI4B (GmCHI4s) in soybean. The real-time quantitative polymerase chain reaction showed that GmCHI4s were expressed primarily in soybean root, but were also present in other tissues, including the stem, leaf, and seed with a low expression level. Overexpression of GmCHI4s was able to significantly improve some beneficial traits of the transformed hair roots of cotyledon or composite plants under salt stress conditions. Root length, root wet weight, and the underground biomass was increased, and the elevation of MDA content was inhibited under 100 mmol L⁻¹ or 150 mmol L⁻¹ NaCl treatment. Leaf chlorophyll content was elevated in overexpressed GmCHI4A composite plants under 150 mmol L⁻¹ NaCl treatment. The expression levels of salt-stress-related genes GmSOD1, GmAPX1, GmSOS1, and GmNHX1 were significantly upregulated in overexpressed GmCHI4 hairy roots compared to that in empty-vector-expressed hairy roots. The above results indicated GmCHI4s’ potential action against salt stress. Furthermore, overexpression of GmCHI4A and GmCHI4B increased the total isoflavone content by six times and three times, respectively. Glycitin and glycitein levels were significantly elevated in the overexpressed GmCHI4A hairy roots, while glycitin, genistin, daidzein, and genistein were significantly increased in overexpressed GmCHI4B hairy roots. This study identified a new function of the CHI-like gene, as well as providing a new selected gene for salt tolerance and isoflavone improvement using biotechnological approaches in soybean. Full article

Citrus is an important type of fruit, with antioxidant bioactivity. However, the variations in the antioxidant ability of different tissues in citrus and its metabolic and molecular basis remain unclear. Here, we assessed the antioxidant capacities of 12 tissues from Citrus reticulata ‘Ponkan’, finding that young leaves and root exhibited the strongest antioxidant capacity. Secondary metabolites accumulated differentially in parts of the citrus plant, of which flavonoids were enriched in stem, leaf, and flavedo; phenolic acids were enriched in the albedo, while coumarins were enriched in the root, potentially explaining the higher antioxidant capacities of these tissues. The spatially specific accumulation of metabolites was related to the expression levels of biosynthesis-related genes such as chalcone synthase (CHS), chalcone isomerase (CHI), flavone synthase (FNS), O-methyltransferase (OMT), flavonoid-3′-hydroxylase (F3′H), flavonoid-6/8-hydroxylase (F6/8H), p-coumaroyl CoA 2′-hydroxylase (C2′H), and prenyltransferase (PT), among others, in the phenylpropane pathway. Weighted gene co-expression network analysis (WGCNA) identified modules associated with flavonoids and coumarin content, among which we identified an OMT involved in coumarin O-methylation, and related transcription factors were predicted. Our study identifies key genes and metabolites influencing the antioxidant capacity of citrus, which could contribute to the enhanced understanding and utilization of bioactive citrus components. Full article

Peanut Fusarium rot, which is widely observed in the main peanut-producing areas in China, has become a significant factor that has limited the yield and quality in recent years. It is highly urgent and significant to clarify the regulatory mechanism of peanuts in response to Fusarium oxysporum. In this study, transcriptome and proteome profiling were combined to provide new insights into the molecular mechanisms of peanut stems after F. oxysporums infection. A total of 3746 differentially expressed genes (DEGs) and 305 differentially expressed proteins (DEPs) were screened. The upregulated DEGs and DEPs were primarily enriched in flavonoid biosynthesis, circadian rhythm-plant, and plant–pathogen interaction pathways. Then, qRT-PCR analysis revealed that the expression levels of phenylalanine ammonia-lyase (PAL), chalcone isomerase (CHI), and cinnamic acid-4-hydroxylase (C4H) genes increased after F. oxysporums infection. Moreover, the expressions of these genes varied in different peanut tissues. All the results revealed that many metabolic pathways in peanut were activated by improving key gene expressions and the contents of key enzymes, which play critical roles in preventing fungi infection. Importantly, this research provides the foundation of biological and chemical analysis for peanut disease resistance mechanisms. Full article