Molecular evidence of phenylpropanoid and flavonoid biosynthesis modulation for copper oxide nanoparticles exposure in water spinach

BackgroundWater spinach (Ipomoea aquatica) is an important heat-resistant leafy vegetable that can survive under long-time heat stress condition. However, the physiological characteristics and molecular changes in its response to heat stress are poorly understood.ResultsIn this study the selected water spinach cultivars with different thermo resistance and their physiological response to heat stress were examined. Under prolonged heat stress, plant growth was inhibited in all tested cultivars. This inhibition was accompanied by the reduction of photosynthetic performance. The reactive oxygen species system in terms of superoxide and hydrogen peroxide contents, as well as antioxidant polyphenols, were evaluated. The results showed that prolonged heat stress caused reduced antioxidant capacity, but the role of antioxidant capacity in a prolonged thermotolerance was not predominant. Transcriptomic analysis of the water spinach subjected to heat stress revealed that 4145 transcripts were specifically expressed with 2420 up-regulated and 1725 down-regulated in heat-sensitive and heat-tolerant cultivars treated with 42 °C for 15 days. Enrichment analysis of these differentially expressed genes showed that the main metabolic differences between heat-sensitive and heat-tolerant cultivars were the carbohydrate metabolism and phenylpropanoid biosynthesis. The results of carbohydrate profiles and RT-qPCR also suggested that heat stress altered carbohydrate metabolism and associated changes in transcriptional level of genes involved in sugar transport and metabolic transition.ConclusionsThe prolonged heat stress resulted in a reduced antioxidant capacity while the role of antioxidant capacity in a prolonged thermotolerance of water spinach was not predominant. Transcriptome analysis and the measurement of carbohydrates as well as the gene expression evaluation indicated that the response of the metabolic pathway such as carbohydrate and phenylpropanoid biosynthesis to heat stress may be a key player in thermo resistance.

Field blanching is a process used in agriculture to obtain sweet, delicious, and tender stems of water dropwort by obstructing sunlight. The nutritional and transcriptomic profiling of blanched water dropwort has been investigated in our previous studies. However, the effect of blanching on the production of secondary metabolites and different vitamins in water dropwort has not been investigated at the transcriptomic level. This study explored the transcriptomic variations in the phenylpropanoid biosynthesis, flavonoid biosynthesis, and different vitamin biosynthesis pathways under different blanching periods in the water dropwort stems (pre-blanching, mid-blanching, post-blanching, and control). The results show that polyphenol and flavonoid contents decreased; however, the contents of vitamins (A, B1, B2, and C) and antioxidant activity increased significantly after blanching. Furthermore, the Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis of blanched water dropwort showed the downregulation of many important genes involved in phenylpropanoid and flavonoid biosynthesis pathways, and the downregulation of these genes might be the reason for the reduction in polyphenol and flavonoid contents. We also examined and highlighted the genes involved in the higher vitamin content, antioxidant activity, pale color, tenderness, and sweetness of the blanched stem of water dropwort. In conclusion, the present study explored the role of phenylpropanoid and vitamin biosynthesis, and it will provide a basis for future investigation and application in the blanch cultivation of water dropwort.

Somatic embryogenesis (SE) techniques have been established for micropropagation or basic research related to plant development in many conifer species. The frequent occurrence of non-embryogenic callus (NEC) during SE has impose constraints on the application of somatic embryogenesis SE in Larix kaempferi (Lamb.) Carr, but the potential regulatory mechanisms are poorly understood. In this study, integrated transcriptomic and metabolomic analyses were performed in embryogenic callus (EC) and NEC originating from a single immature zygotic embryo to better decipher the key molecular and metabolic mechanisms required for embryogenic potential maintenance. The results showed that a total of 13,842 differentially expressed genes (DEGs) were found in EC and NEC, among which many were enriched in plant hormone signal transduction, starch and sucrose metabolism, phenylpropanoid biosynthesis, flavonoid biosynthesis, and the biosynthesis of amino acids pathways. Metabolite profiling showed that 441 differentially accumulated metabolites (DAMs) were identified in EC and NEC. Both EC and NEC had vigorous primary metabolic activities, while most secondary metabolites were upregulated in NEC. Many totipotency-related transcription factor (TF) genes such as BBMs, WUSs, and LEC1 showed higher expression levels in EC compared with NEC, which may result in the higher accumulation of indole 3-acetic acid (IAA) in EC. NEC was characterized by upregulation of genes and metabolites associated with stress responses, such as DEGs involved in jasmonic acid (JA) and ethylene (ETH) biosynthesis and signal transduction pathways, and DEGs and DAMs related to phenylpropanoid and flavonoid biosynthesis. We predicted and analyzed TFs that could target several key co-expressed structural DEGs including two C4H genes, two CcoAOMT genes and three HCT genes involved in phenylpropanoid and flavonoid biosynthesis. Based on the targeted relationship and the co-expression network, two ERFs (Lk23436 and Lk458687), one MYB (Lk34626) and one C2C2-dof (Lk37167) may play an important role in regulating phenolic acid and flavonoid biosynthesis by transcriptionally regulating the expression of these structural genes. This study shows an approach involving integrated transcriptomic and metabolic analyses to obtain insights into molecular events underlying embryogenic potential maintenance and the biosynthesis mechanisms of key metabolites involving TF regulation, which provides valuable information for the improvement of SE efficiency in L. kaempferi.

Aphid, Aphis gossypii Glover, is a pest that significantly affects cucumbers (Cucumis sativus L.). Phloem protein 2 (PP2) is a conserved phloem lectin. Our previous study showed that the expression of CsPP2-A1 under aphid attack affected the accumulation of flavonoids and total phenolics in cucumber. The novel mechanism of lectin CsPP2-A1 mediating secondary metabolites affecting aphid resistance in cucumbers needs to be investigated. The weight and length of aphids on CsPP2-A1 overexpression (CsPP2-A1-OE) cucumber plants significantly reduced compared to wild-type (WT). Conversely, aphids on CsPP2-A1 RNA interference (CsPP2-A1-RNAi) plants showed the opposite trend. Using secondary metabolomics, small molecular weight secondary metabolites were qualitatively and quantitatively assessed in WT and transgenic cucumber plants after aphid inoculation. The overexpression of CsPP2-A1 resulted in the up-regulation of differential metabolites (DMs) in phenylpropanoid biosynthesis, whereas interference expression of CsPP2-A1 led to a down-regulation of DMs in the flavonoid biosynthesis. Concurrently, it was observed that the CAD activity and the expression of the CsPAL, and CsCAD in OE-2 were up-regulated significantly. A significant reduction in the activities of CHI, F3H, and the expression of CsF3H, CsCHS, CsFLS, and CsCCR was noted in RNAi-2. CsPP2-A1 indirectly affects the growth and development of aphids via mediation of phenylpropanoid and flavonoid biosynthesis. The indirect effects of the interaction of CsPP2-A1 with aphids offer insights into plant-insect interaction studies. © 2025 Society of Chemical Industry.

Suaeda glauca Bunge produces dimorphic seeds on the same plant, with brown seeds displaying non-dormant characteristics and black seeds exhibiting intermediate physiological dormancy traits. Previous studies have shown that black seeds have a very low germination rate under natural conditions, but exogenous GA3 effectively enhanced the germination rate of black seeds. However, the physiological and molecular mechanisms underlying the effects of GA3 on S. glauca black seeds are still unclear. In this study, transcriptomic profiles of seeds at different germination stages with and without GA3 treatment were analyzed and compared, and the TTF, H2O2, O2 -, starch, and soluble sugar contents of the corresponding seed samples were determined. The results indicated that exogenous GA3 treatment significantly increased seed vigor, H2O2, and O2 - contents but decreased starch and soluble sugar contents of S. glauca black seeds during seed dormancy release. RNA-seq results showed that a total of 1136 DEGs were identified in three comparison groups and were involved mainly in plant hormone signal transduction, diterpenoid biosynthesis, flavonoid biosynthesis, phenylpropanoid biosynthesis, and carbohydrate metabolism pathway. Among them, the DEGs related to diterpenoid biosynthesis (SgGA3ox1, SgKAO and SgGA2ox8) and ABA signal transduction (SgPP2Cs) could play important roles during seed dormancy release. Most genes involved in phenylpropanoid biosynthesis were activated under GA3 treatment conditions, especially many SgPER genes encoding peroxidase. In addition, exogenous GA3 treatment also significantly enhanced the expression of genes involved in flavonoid synthesis, which might be beneficial to seed dormancy release. In accordance with the decline in starch and soluble sugar contents, 15 genes involved in carbohydrate metabolism were significantly up-regulated during GA3-induced dormancy release, such as SgBAM, SgHXK2, and SgAGLU, etc. In a word, exogenous GA3 effectively increased the germination rate and seed vigor of S. glauca black seeds by mediating the metabolic process or signal transduction of plant hormones, phenylpropanoid and flavonoid biosynthesis, and carbohydrate metabolism processes. Our results provide novel insights into the transcriptional regulation mechanism of exogenous GA3 on the dormancy release of S. glauca black seeds. The candidate genes identified in this study may be further studied and used to enrich our knowledge of seed dormancy and germination.

Ginkgo leaves are always resources for flavonoids pharmaceutical industry. However, the effect of the elevation and tree age changes on flavonoid biosynthesis have not been detailly explored in Ginkgo leaves. In addition, whether these environmental pressures have similar effects on the biosynthesis of other non-flavonoids polyphenolics in phenylpropanoid biosynthesis is not known at present. In this research, de novo transcriptome sequencing of Ginkgo leaves was performed coupled with ultra-performance liquid chromatography/quadrupole time-of-flight mass spectrometry analyses to obtain a comprehensive understanding of the influence of elevation and tree age on phenylpropanoid biosynthesis. A total of 557,659,530 clean reads were assembled into 188,155 unigenes, of which 135,102 (71.80%) were successfully annotated in seven public databases. The putative DFRs, LARs, and ANRs were significantly up-regulated with the increase of elevation in young Ginkgo tree leaves. The relative concentration of flavonoid derivatives with high parent ion intensity was likely to imply that the elevation increase promoted the biosynthesis of flavonoids. Complex gene variations involved in flavonoid biosynthesis were observed with the tree age increase. However, flavonoid derivatives analysis predicted that the rise of tree age was more likely to be detrimental to the flavonoids manufacture. Otherwise, multiple genes implicated in the synthesis of hydroxycinnamates, lignin, and lignan exhibited fluctuations with the elevation increase. Significantly up-regulated CADs and down-regulated PRDs potentially led to the accumulation of p-Coumaryl alcohol, one of the lignin monomers, and might inhibit further lignification. Overall, the putative DFRs seemed to show more considerable variability toward these stress, and appeared to be the main regulatory point in the flavonoid biosynthesis. Light enhancement caused by elevation increase may be the main reason for flavonoids accumulation. Flavonoid biosynthesis exhibited a greater degree of perturbation than that of hydroxycinnamates, lignins and lignans, potentially suggesting that flavonoid biosynthesis might be more susceptible than other branch pathways involved in phenylpropanoid biosynthesis. This research effectively expanded the functional genomic library and provide new insights into phenylpropanoid biosynthesis in Ginkgo.

The somatic embryogenesis (SE) process of plants is regulated by exogenous hormones. During the SE, different genes sensitively respond to hormone signals through complex regulatory networks to exhibit plant totipotency. When cultured in indole-3-butyric acid (IBA) concentration gradient medium supplemented with 0 mg dm−3, 0.025 mg dm−3, and 0.05 mg dm−3 IBA, the callus differentiation rate first increased then decreased in cotton. To characterize the molecular basis of IBA-induced regulating SE, transcriptome analysis was conducted on embryogenic redifferentiation. Upon the examination of the IBA’s embryogenic inductive effect, it was revealed that pathways related to plant hormone signal transduction and alcohol degradation were significantly enriched in the embryogenic responsive stage (5 days). The photosynthesis, alcohol metabolism and cell cycle pathways were specifically regulated in the pre-embryonic initial period (20 days). Upon the effect of the IBA dose, in the embryogenic responsive stage (5 days), the metabolism of xenobiotics by the cytochrome P450 pathway and secondary metabolism pathways of steroid, flavonoid, and anthocyanin biosynthesis were significantly enriched. The phenylpropanoid, brassinosteroid, and anthocyanin biosynthesis pathways were specifically associated in the pre-embryonic initial period (20 days). At different developmental stages of embryogenic induction, photosynthesis, flavonoid biosynthesis, phenylpropanoid biosynthesis, mitogen-activated protein kinase (MAPK) signaling, xenobiotics metabolism by cytochrome P450, and brassinosteroid biosynthesis pathways were enriched at low a IBA concentration. Meanwhile, at high IBA concentration, the carbon metabolism, alcohol degradation, circadian rhythm and biosynthesis of amino acids pathways were significantly enriched. The results reveal that complex regulating pathways participate in the process of IBA-induced redifferentiation in cotton somatic embryogenesis. In addition, collections of potential essential signaling and regulatory genes responsible for dose IBA-induced efficient embryogenic redifferentiation were identified. Quantitative real-time PCR (qRT-PCR) was performed on the candidate genes with different expression patterns, and the results are basically consistent with the RNA-seq data. The results suggest that the complicated and concerted IBA-induced mechanisms involving multiple cellular pathways are responsible for dose-dependent plant growth regulator-induced SE. This report represents a systematic study and provides new insight into molecular signaling and regulatory basis underlying the process of dose IBA-induced embryogenic redifferentiation during SE.

Saposhnikovia divaricata, obtained from the desiccated roots of the plant Saposhnikovia divaricata (Turcz.) Schischk, is a medicinal material with its phloem known to be richer in chromones than its xylem. To understand this disparity, we compared the xylem and phloem using high-performance liquid chromatography (HPLC), transcriptomics, and nontargeted metabolomics. HPLC revealed that cimifugin was six times higher in the phloem. Conversely, prim-O-glucosylcimifugin and 4'-O-β-D-glucosyl-5-O-methylvisamminol were 1.2 and 1.6 times more abundant in the xylem, respectively (p < 0.01). Transcriptome sequencing identified nine differentially expressed genes, predominantly involved in phenylpropanoid biosynthesis (PAL, HCT, CCR, CSE, COMT, and CCoAOMT). Nontargeted metabolomics identified 229 differential metabolites, also largely enriched in phenylpropanoid biosynthesis. Integrated analysis revealed a strong positive correlation between the gene COMT and two metabolites (trans-cinnamaldehyde and myricatomentoside I), identifying COMT as a key candidate gene governing quality. Our findings demonstrate that phenylpropanoid, flavonoid, and lignin biosynthesis pathways crucially impact secondary metabolite accumulation in Saposhnikovia divaricata, providing new insights for its exploitation.

Rhododendron is an important woody ornamental plant, and breeding varieties with different colors is a key research goal. Although there have been a few reports on the molecular mechanisms of flower colors and color patterning in Rhododendron, it is still largely unknown what factors regulate flower pigmentation in Rhododendron. In this study, the flower color variation cultivar 'Yanzhi Mi' and the wild-type (WT) cultivar 'Dayuanyangjin' were used as research objects, and the pigments and transcriptomes of their petals during five flower development stages were analyzed and compared. The results showed that derivatives of cyanidin, peonidin and pelargonidin might be responsible for the pink color of mutant petals and that the S2 stage was the key stage of flower color formation. In total, 412,910 transcripts and 2780 differentially expressed genes (DEGs) were identified in pairwise comparisons of WT and mutant petals. GO and KEGG enrichment analyses of the DEGs showed that 'DNA-binding transcription factor activity', 'Flavonoid biosynthesis' and 'Phenylpropanoid biosynthesis' were more active in mutant petals. Early anthocyanin pathway candidate DEGs (CHS3-CHS6, CHI, F3Hs and F3'H) were significantly correlated and were more highly expressed in mutant petals than in WT petals in the S2 stage. An R2R3-MYB unigene (TRINITY_DN55156_c1_g2) was upregulated approximately 10.5-fold in 'Yanzhi Mi' petals relative to 'Dayuanyangjin' petals in the S2 stage, and an R2R3-MYB unigene (TRINITY_DN59015_c3_g2) that was significantly downregulated in 'Yanzhi Mi' petals in the S2 stage was found to be closely related to Tca MYB112 in cacao. Taken together, the results of the present study could shed light on the molecular basis of anthocyanin biosynthesis in two Rhododendron obtusum cultivars and may provide a genetic resource for breeding varieties with different flower colors.

BackgroundNotopterygium incisum seeds have both morphological and physiological dormancy characteristics and require stratification to break seed dormancy, but the mechanism of seed dormancy release during stratification is still unclear. In this study, different stages of N. incisum seed stratification were employed as experimenta objects, and the dynamic changes during seed dormancy release were studied through embryo morphology, physiological index determination, transcriptome, and metabolome.Results(1) Stratification treatment reduced the content of stored nutrients in N. incisum seeds, significantly changed enzyme activity, reduced ABA content, and increased GA3 and IAA contents. (2) A total of 110,539 differentially expressed genes (DEGs) and 1656 metabolites (DAMs) were identified during dormancy release. Transcriptome analysis showed that after the dormancy of N. incisum seeds was released, the expression of genes in the abscisic acid signaling pathway (ABI1, PP2CA, ABI5 and ABF4) and the gibberellin signaling pathway (GAI, GAI1 and RGL1) were significantly down-regulated, and there were significant changes in the differentially expressed genes in the auxin, cytokinin and ethylene signaling pathways. The genes related to starch and sucrose metabolism were up-regulated during dormancy release. The genes related to phenylpropanoid and flavonoid biosynthesis were significantly up-regulated after dormancy release. (3) Combined transcriptomics and metabolomics analysis showed that phenylpropanoid biosynthesis and flavonoid biosynthesis are the key pathways for the dormancy release of N. incisum seeds. (4) Metabolomics analysis showed that the accumulation of metabolites of the phenylpropanoid biosynthesis pathway (p-coumaric acid, coniferyl aldehyde, coniferyl glycoside, 5-caffeoylshikimic acid and sinapinic acid) decreased during and after the dormancy release of N. incisum seeds, while the accumulation of flavonoids such as quercetin, rutin, delphinidin and naringenin chalcone increased significantly after dormancy release.ConclusionDormancy release in N. incisum seeds involves differential regulation of hormones, carbohydrates, phenylpropanoids, and flavonoid metabolites. Our results provide important insights into the molecular regulatory network of dormancy release in N. incisum seeds.Supplementary InformationThe online version contains supplementary material available at 10.1186/s12864-025-12021-x.

A total of 299,113 unigenes were generated and 15,817 DEGs were identified. We identified candidate genes associated with the regulation of catechins biosynthesis during leaf development in tea plant. The tea plant (Camellia sinensis (L.) O. Kuntze) is one of the most economically significant crops worldwide because of its positive effects on human health. The health benefits of tea are mainly attributed to catechins, which are the predominant polyphenols that accumulate in tea. Catechins are products of the phenylpropanoid and flavonoid biosynthetic pathways. Although catechins were identified in tea leaves long ago, the molecular mechanisms regulating catechins biosynthesis remain unclear. To identify candidate genes involved in catechins biosynthesis, we analyzed the transcriptomes of tea leaves during five different leaf stages of development using RNA-seq. Approximately 809 million high-quality reads were obtained, trimmed, and assembled into 299,113 unigenes with an average length of 565bp. A total of 15,817 unigenes were differentially expressed during the different stages of leaf development. These differentially expressed genes were enriched in a variety of processes such as the regulation of the cell cycle, starch and sucrose metabolism, photosynthesis, phenylpropanoid biosynthesis, phenylalanine metabolism, and flavonoid biosynthesis. Based on their annotations, 51 of these differentially expressed unigenes are involved in phenylpropanoid and flavonoid biosynthesis. Furthermore, transcription factors such as MYB, bHLH and MADS, which may involve in the regulation of catechins biosynthesis, were identified through co-expression analysis of transcription factors and structural genes. Real-time PCR analysis of candidate genes indicated a good correlation with the transcriptome data. These findings increase our understanding of the molecular mechanisms regulating catechins biosynthesis in the tea plant.

Metabolomics integrated with transcriptomics provides insights into the phenylpropanoids biosynthesis pathway in Lilium davidii var. unicolor and L. lancifolium Thunb.

Transcriptomic and metabolomic perspectives for the growth of alfalfa (Medicago sativa L.) seedlings with the effect of vanadium exposure

Metabolic composition can have potential impact on several vital agronomic traits, and metabolomics, which represents the bioactive compounds in plant tissues, is widely considered as a powerful approach for linking phenotype–genotype interactions. However, metabolites related to cane traits such as sugar content, rind color, and texture differences in different sugarcane cultivars using metabolome integrated with transcriptome remain largely inconclusive. In this study, metabolome integrated with transcriptome analyses were performed to identify and quantify metabolites composition, and have better insight into the molecular mechanisms underpinning the different cane traits, namely, brix, rind color, and textures in the stems (S) and leaves (L) of sugarcane varieties FN41 and 165402. We also identified metabolites and associated genes in the phenylpropanoid and flavonoid biosynthesis pathways, starch and sucrose metabolism. A total of 512 metabolites from 11 classes, with the vast majority (122) belonging to flavonoids were identified. Moreover, the relatively high amount of D-fructose 6-p, D-glucose6-p and glucose1-p detected in FN41L may have been transported and distributed by source and sink of the cane, and a majority of them reached the stem of sugarcane FN41L, thereby promoting the high accumulation of sugar in FN41S. Observations also revealed that genes such as C4H, CHS, F3H, F3’H, DFR, and FG2 in phenylpropanoid and flavonoid biosynthesis pathways were the major factors impacting the rind color and contrasting texture of FN41 and 165204. Further analysis revealed that weighted gene co-expression network analysis (WGCNA) hub genes and six transcription factors, namely, Tify and NAC, MYB-related, C2C2-Dof, WRKY, and bHLH play a key role in phenylpropanoid biosynthesis, flavone and flavonol biosynthesis, starch and sucrose metabolism. Additionally, metabolites such as L-phenylalanine, tyrosine, sinapaldehyde, pinobanksin, kaempferin, and nictoflorin were the potential drivers of phenotypic differences. Our finding also demonstrated that genes and metabolites in the starch and sucrose metabolism had a significant effect on cane sugar content. Overall, this study provided valuable insight into the molecular mechanisms underpinning high sugar accumulation and rind color in sugarcane, which we believe is important for future sugarcane breeding programs and the selection of high biomass varieties.

De novo transcriptome assembly and metabolomic analysis of three tissue types in Cinnamomum cassia