RESUMEN
Scopoletin and chlorogenic acid (CGA) are important polyphenols that regulate plant growth, development, and stress resistance. The ERF transcription factor WAX INDUCER1 (WIN1) promotes the biosynthesis of cutin, suberine, and wax. However, its full roles in regulating the accumulation of plant secondary metabolites still remain to be further clarified. In this study, NtWIN1 gene encoding a SHINE-type AP2/ERF transcription factor of the Va subgroup was identified from N. tabacum. NtWIN1 showed high expression levels in tobacco stems, sepals, and pistils. Overexpression (OE) and knock-out of NtWIN1 showed that it promoted the accumulation of total polyphenols and altered their composition. Compare to that of WT plants, the CGA contents significantly increased by 25%-50% in the leaves, flowers, and capsules of OE lines, while the scopoletin contents in the OE plants significantly decreased by 30%-67%. In contrast, the CGA contents in ntwin1 lines reduced by 23%-26%, and the scopoletin contents in ntwin1 increased by 38%-75% compare to that of WT plants. Chromatin immunoprecipitation and Dual-Luc transcription activation assays showed that NtWIN1 could bind to the promoters of NtF6'H1 and NtCCoAMT, thereby modulating their expression. The scopoletin content in ntwin1/ntf6'h1 double mutant was significantly lower than that in ntwin1 and WT plants, but showed no significant differences with that in ntf6'h1 mutant, further indicating that the inhibition of NtWIN1 on scopoletin accumulation depends on the activity of NtF6'H1. Our study illustrates the new roles of NtWIN1, and provides a possible target for regulating the synthesis of polyphenols in tobacco.
Asunto(s)
Ácido Clorogénico , Regulación de la Expresión Génica de las Plantas , Nicotiana , Proteínas de Plantas , Escopoletina , Nicotiana/genética , Nicotiana/metabolismo , Escopoletina/metabolismo , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Ácido Clorogénico/metabolismo , Factores de Transcripción/metabolismo , Factores de Transcripción/genética , Metionina Adenosiltransferasa/metabolismo , Metionina Adenosiltransferasa/genética , Plantas Modificadas Genéticamente/metabolismo , Plantas Modificadas Genéticamente/genética , Genes de PlantasRESUMEN
Alternaria leaf blight (ALB), caused by a necrotrophic fungus Alternaria brassicae is a serious disease of oleiferous Brassicas resulting in significant yield losses worldwide. No robust resistance against A. brassicae has been identified in the Brassicas. Natural accessions of Arabidopsis show a spectrum of responses to A. brassicae ranging from high susceptibility to complete resistance. To understand the molecular mechanisms of resistance/ susceptibility, we analysed the comparative changes in the transcriptome profile of Arabidopsis accessions with contrasting responses- at different time points post-infection. Differential gene expression, GO enrichment, pathway enrichment, and weighted gene co-expression network analysis (WGCNA) revealed reprogramming of phenylpropanoid biosynthetic pathway involving lignin, hydroxycinnamic acids, scopoletin, anthocyanin genes to be highly associated with resistance against A. brassicae. T-DNA insertion mutants deficient in the biosynthesis of coumarin scopoletin exhibited enhanced susceptibility to A. brassicae. The supplementation of scopoletin to medium or exogenous application resulted in a significant reduction in the A. brassicae growth. Our study provides new insights into the transcriptome dynamics in A. brassicae-challenged Arabidopsis and demonstrates the involvement of coumarins in plant immunity against the Brassica pathogen A. brassicae.
Asunto(s)
Alternaria , Arabidopsis , Resistencia a la Enfermedad , Regulación de la Expresión Génica de las Plantas , Enfermedades de las Plantas , Transcriptoma , Arabidopsis/genética , Arabidopsis/microbiología , Arabidopsis/inmunología , Alternaria/fisiología , Enfermedades de las Plantas/microbiología , Enfermedades de las Plantas/genética , Enfermedades de las Plantas/inmunología , Resistencia a la Enfermedad/genética , Escopoletina/metabolismo , Perfilación de la Expresión Génica , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismoRESUMEN
OBJECTIVES: The hepatoprotective properties of scopoletin have been explored in carbon tetrachloride (CCl4) induced liver injury but not in drug-induced liver injury (DILI) scenarios. Only N-acetyl-cysteine (NAC) has proven efficacy in DILI treatment. Accordingly, we conducted a study to assess the hepatoprotective action of scopoletin in the anti-tubercular treatment (ATT)-DILI model in Wistar rats, if any. METHODS: A total of 36 rats were evaluated, with six in each group. A 36-day ATT at 100â¯mg/kg dose for isoniazid, 300â¯mg/kg for rifampicin and 700â¯mg/kg for pyrazinamide were fed to induce hepatotoxicity in rats. Group I and II-VI received normal saline and ATT, respectively. Oral scopoletin (1,5 and 10â¯mg/kg) and NAC 150â¯mg/kg were administered in groups III, IV, V and VI, respectively, once daily for the last 15 days of the experiment. LFT monitoring was performed at baseline, days 21, 28, and 36. Rats were sacrificed for the histopathology examination. RESULTS: Aspartate transaminase (AST), alanine transaminase (ALT), alkaline phosphatase (ALP) and bilirubin levels were significantly increased in group II (receiving ATT) compared to normal control on day 28 and day 36 (p<0.05). All three doses of scopoletin and NAC groups led to the resolution of AST, ALT, ALP, and bilirubin changes induced by ATT medications effect beginning by day 28 and persisting on day 36 (p<0.01). An insignificant effect was observed on albumin and total protein levels. The effect was confirmed with antioxidants and histopathology analysis. CONCLUSIONS: The study confirms the hepatoprotective efficacy of scopoletin in a more robust commonly encountered liver injury etiology.
Asunto(s)
Enfermedad Hepática Inducida por Sustancias y Drogas , Escopoletina , Ratas , Animales , Ratas Wistar , Escopoletina/farmacología , Escopoletina/uso terapéutico , Escopoletina/metabolismo , Extractos Vegetales/farmacología , Extractos Vegetales/uso terapéutico , Antituberculosos/toxicidad , Enfermedad Hepática Inducida por Sustancias y Drogas/tratamiento farmacológico , Hígado , Bilirrubina/metabolismo , Fosfatasa Alcalina/metabolismo , Tetracloruro de Carbono/metabolismo , Tetracloruro de Carbono/farmacología , Alanina Transaminasa/metabolismoRESUMEN
This study investigated whether scopoletin could protect INS-1 pancreatic ß cells from apoptosis and oxidative stress caused by high glucose. Cells were pretreated with glucose (5.5 or 30 mM) and then treated with 0, 5, 10, 25, or 50 µM Scopoletin. Cell viability and insulin secretion were measured in addition to ROS, TBARS, NO and antioxidant enzymes. Western blot analysis and flow cytometric assessment of apoptosis were also carried out. High glucose of 30 mM caused glucotoxicity and cell death in INS-1 pancreatic ß cells. However, 5, 10, 25 or 50 µM scopoletin increased the level of cell viability as concentrations increased. The levels of ROS, TBARS, and NO increased by high glucose were significantly decreased after scopoletin treatment. Scopoletin also raised antioxidant enzyme activities up against oxidative stress produced by high glucose. These effects influenced the apoptosis pathway, raising levels of anti-apoptotic protein, Bcl-2, and reducing levels of pro-apoptotic proteins, including JNK, Bax, cytochrome C, and caspase 9. Annexin V/propidium staining indicated that scopoletin significantly lowered high glucose-produced apoptosis. These results indicate that scopoletin can protect INS-1 pancreatic ß cells from glucotoxicity caused by high glucose and have potential as a pharmaceutical material to protect the pancreatic ß cells.
Asunto(s)
Células Secretoras de Insulina , Células Secretoras de Insulina/metabolismo , Escopoletina/farmacología , Escopoletina/metabolismo , Antioxidantes/metabolismo , Especies Reactivas de Oxígeno/metabolismo , Sustancias Reactivas al Ácido Tiobarbitúrico/metabolismo , Apoptosis , Estrés Oxidativo , Glucosa/toxicidad , Glucosa/metabolismo , Insulina/metabolismoRESUMEN
Coumarins can fight pathogens and are thus promising for crop protection. Their biosynthesis, however, has not yet been engineered in crops. We tailored the constitutive accumulation of coumarins in transgenic Nicotiana benthamiana, Glycine max and Arabidopsis thaliana plants, as well as in Nicotiana tabacum BY-2 suspension cells. We did so by overexpressing A. thaliana feruloyl-CoA 6-hydroxylase 1 (AtF6'H1), encoding the key enzyme of scopoletin biosynthesis. Besides scopoletin and its glucoside scopolin, esculin at low level was the only other coumarin detected in transgenic cells. Mechanical damage of scopolin-accumulating tissue led to a swift release of scopoletin, presumably from the scopolin pool. High scopolin levels in A. thaliana roots coincided with reduced susceptibility to the root-parasitic nematode Heterodera schachtii. In addition, transgenic soybean plants were more tolerant to the soil-borne pathogenic fungus Fusarium virguliforme. Because mycotoxin-induced accumulation of reactive oxygen species and cell death were reduced in the AtF6'H1-overexpressors, the weaker sensitivity to F. virguliforme may be caused by attenuated oxidative damage of coumarin-hyperaccumulating cells. Together, engineered coumarin accumulation is promising for enhanced disease resilience of crops.
Asunto(s)
Arabidopsis , Micotoxinas , Arabidopsis/metabolismo , Escopoletina/metabolismo , Micotoxinas/metabolismo , Susceptibilidad a Enfermedades/metabolismo , Cumarinas/metabolismo , Estrés Oxidativo , Raíces de Plantas/genética , Raíces de Plantas/metabolismoRESUMEN
Although mainly known for producing artemisinin, Artemisia annua is enriched in phenylpropanoid glucosides (PGs) with significant bioactivities. However, the biosynthesis of A. annua PGs is insufficiently investigated. Different A. annua ecotypes from distinct growing environments accumulate varying amounts of metabolites, including artemisinin and PGs such as scopolin. UDP-glucose:phenylpropanoid glucosyltransferases (UGTs) transfers glucose from UDP-glucose in PG biosynthesis. Here, we found that the low-artemisinin ecotype GS produces a higher amount of scopolin, compared to the high-artemisinin ecotype HN. By combining transcriptome and proteome analyses, we selected 28 candidate AaUGTs from 177 annotated AaUGTs. Using AlphaFold structural prediction and molecular docking, we determined the binding affinities of 16 AaUGTs. Seven of the AaUGTs enzymatically glycosylated phenylpropanoids. AaUGT25 converted scopoletin to scopolin and esculetin to esculin. The lack of accumulation of esculin in the leaf and the high catalytic efficiency of AaUGT25 on esculetin suggest that esculetin is methylated to scopoletin, the precursor of scopolin. We also discovered that AaOMT1, a previously uncharacterized O-methyltransferase, converts esculetin to scopoletin, suggesting an alternative route for producing scopoletin, which contributes to the high-level accumulation of scopolin in A. annua leaves. AaUGT1 and AaUGT25 responded to induction of stress-related phytohormones, implying the involvement of PGs in stress responses.
Asunto(s)
Artemisia annua , Artemisininas , Artemisia annua/metabolismo , Escopoletina/química , Escopoletina/metabolismo , Escopoletina/farmacología , Esculina/metabolismo , Multiómica , Simulación del Acoplamiento Molecular , Artemisininas/metabolismo , Glucósidos/metabolismo , Glucosa/metabolismo , Uridina Difosfato/metabolismoRESUMEN
Cassava landraces are impacted by post-harvest physiological deterioration (PPD). 34 primary/secondary metabolites (carotenes, flavonols, indols, phenolic, hydroxycinnamic, and organic acids) were analysed using HPLC/GC-MS in 72 landraces harvested 8 months after planting (MAP) to clarify whether these compounds may play a role in PPD tolerance. Cluster analysis differentiated a first group with high organic acids contents, citric acid being dominant, a second group with landraces high in tryptophan, a third group including landraces with high phenolic and hydroxycinnamic acids content, and a fourth group characterised by 8 carotenoids. PPD tolerant and susceptible landraces were present in each group. To determine if PPD is related to age of harvest, 174 landraces were harvested at 6, 8, 10 and 12 MAP. Scopoletin, sucrose and glucose were analysed. PPD was positively correlated with DMC and negatively correlated with scopoletin at all ages of harvest. Scopoletin is a useful biomarker to characterize landraces.
Asunto(s)
Manihot , Escopoletina , Escopoletina/metabolismo , Manihot/metabolismo , Carotenoides/metabolismo , Verduras/metabolismo , Raíces de Plantas/metabolismoRESUMEN
Coumarins are generally considered to be produced by natural plants. Fungi have been reported to produce coumarins, but their biosynthetic pathways are still unknown. In this study, Fusarium oxysporum GU-7 and GU-60 were isolated from Glycyrrhiza uralensis, and their antioxidant activities were determined to be significantly different. Abundant dipeptide, phenolic acids, and the plant-derived coumarins fraxetin and scopoletin were identified in GU-7 by untargeted metabolomics, and these compounds may account for its stronger antioxidant activity compared to GU-60. Combined with metabolome and RNA sequencing analysis, we identified 24 potentially key genes involved in coumarin biosynthesis and 6 intermediate metabolites. Interestingly, the best hit of S8H, a key gene involved in hydroxylation at the C-8 position of scopoletin to yield fraxetin, belongs to a plant species. Additionally, nondestructive infection of G. uralensis seeds with GU-7 significantly improved the antioxidant activity of seedlings compared to the control group. This antioxidant activity may depend on the biological characteristics of endophytes themselves, as we observed a positive correlation between the antioxidant activity of endophytic fungi and that of their nondestructively infected seedlings. IMPORTANCE Plant-produced coumarins have been shown to play an important role in assembly of the plant microbiomes and iron acquisition. Coumarins can also be produced by some microorganisms. However, studies on coumarin biosynthesis in microorganisms are still lacking. We report for the first time that fraxetin and scopoletin were simultaneously produced by F. oxysporum GU-7 with strong free radical scavenging abilities. Subsequently, we identified intermediate metabolites and key genes in the biosynthesis of these two coumarins. This is the first report on the coumarin biosynthesis pathway in nonplant species, providing new strategies and perspectives for coumarin production and expanding research on new ways for plants to obtain iron.
Asunto(s)
Antioxidantes , Arabidopsis , Antioxidantes/metabolismo , Escopoletina/química , Escopoletina/metabolismo , Arabidopsis/genética , Vías Biosintéticas/genética , Cumarinas/química , Cumarinas/metabolismo , Plantas/metabolismo , Hierro/metabolismoRESUMEN
Uridine diphosphate-dependent glycosyltransferases (UGTs) mediate the glycosylation of plant metabolites, thereby altering their physicochemical properties and bioactivities. Plants possess numerous UGT genes, with the encoded enzymes often glycosylating multiple substrates and some exhibiting substrate inhibition kinetics, but the biological function and molecular basis of these phenomena are not fully understood. The promiscuous monolignol/phytoalexin glycosyltransferase NbUGT72AY1 exhibits substrate inhibition (Ki) at 4 µM scopoletin, whereas the highly homologous monolignol StUGT72AY2 is inhibited at 190 µM. We therefore used hydrogen/deuterium exchange mass spectrometry and structure-based mutational analyses of both proteins and introduced NbUGT72AY1 residues into StUGT72AY2 and vice versa to study promiscuity and substrate inhibition of UGTs. A single F87I and chimeric mutant of NbUGT72AY1 showed significantly reduced scopoletin substrate inhibition, whereas its monolignol glycosylation activity was almost unaffected. Reverse mutations in StUGT72AY2 resulted in increased scopoletin glycosylation, leading to enhanced promiscuity, which was accompanied by substrate inhibition. Studies of 3D structures identified open and closed UGT conformers, allowing visualization of the dynamics of conformational changes that occur during catalysis. Previously postulated substrate access tunnels likely serve as drainage channels. The results suggest a two-site model in which the second substrate molecule binds near the catalytic site and blocks product release. Mutational studies showed that minor changes in amino acid sequence can enhance the promiscuity of the enzyme and add new capabilities such as substrate inhibition without affecting existing functions. The proposed subfunctionalization mechanism of expanded promiscuity may play a role in enzyme evolution and highlights the importance of promiscuous enzymes in providing new functions.
Asunto(s)
Fitoalexinas , Escopoletina , Escopoletina/metabolismo , Glicosilación , Glicosiltransferasas/química , Plantas/metabolismoRESUMEN
Scopoletin is a phenolic coumarin isolated from a variety of plants and was originally used to treat various diseases including arthritis as well as bone-related diseases. The goal of this study was to determine scopoletin's therapeutic potential in an animal model of myocardial infarction induced with ISO. There were five groups of albino male rats. Group I (control) animals were orally treated with olive oil. Group II (scopoletin) animals were pre-treated orally with a 50-mg dosage of scopoletin for 28 days. Group III (ISO-treated) animals were treated with 85 mg/kg of ISO subcutaneously for 2 consecutive days (29th and 30th day). Group IV (scopoletin and ISO) animals were pre-treated orally with 25 mg of scopoletin for 28 days before exposure to ISO. Group V (scopoletin and ISO) animals were pre-treated with 50 mg of scopoletin for 28 days before exposure to ISO. In the ISO-administered animals, a wider heart-to-body weight ratio, a higher heart weight, higher cardiac diagnostic markers, higher MDA levels and related antioxidant levels, inflammatory, and apoptotic markers were observed. Scopoletin pre-treatment with ISO (25 and 50 mg/kg b.wt) significantly reduced heart-to-body weight ratio, cardiac diagnostic markers, MDA, inflammatory markers, and apoptotic markers. Meantime, a pre-treatment with scopoletin increased the levels of antioxidant enzymes. Inflammation and necrosis were observed in the histopathology of heart tissue of ISO-treated animals and these histopathological conditions were reversed by scopoletin pretreatment. The antioxidant and anti-inflammatory properties of ISO-treated rats were shown to be increased by scopoletin, showing its therapeutic potential against cardiovascular diseases. Through the use of its antioxidant and anti-inflammatory properties, scopoletin exhibited anti-myocardial infarction properties. However, further preclinical studies will be required to demonstrate the mechanism of action of scopoletin involved in anti-myocardial infarction.
Asunto(s)
Antioxidantes , Infarto del Miocardio , Ratas , Animales , Isoproterenol/efectos adversos , Isoproterenol/metabolismo , Antioxidantes/metabolismo , Escopoletina/efectos adversos , Escopoletina/metabolismo , Miocardio/metabolismo , Infarto del Miocardio/inducido químicamente , Infarto del Miocardio/tratamiento farmacológico , Antiinflamatorios/farmacología , Peso Corporal , Estrés Oxidativo , Cardiotónicos/efectos adversosRESUMEN
There are abundant polyphenols in tobacco leaves mainly including chlorogenic acid (CGA), rutin, and scopoletin, which not only influence plant growth, development, and environmental adaptation, but also have a great impact on the industrial utilization of tobacco leaves. Few transcription factors regulating the biosynthesis of polyphenols have been identified in tobacco so far. In this study, two NtWRKY33 genes were identified from N. tabacum genome. NtWRKY33a showed higher transcriptional activity than NtWRKY33b, and encoded a nuclear localized protein. Overexpression and knock-out of NtWRKY33a gene revealed that NtWRKY33a inhibited the accumulation of rutin, scopoletin, and total polyphenols, but meanwhile promoted the biosynthesis of CGA. Chromatin immunoprecipitation and Dual-Luc assays indicated that NtWRKY33a could directly bind to the promoters of NtMYB4 and NtHCT, and thus induced the transcription of these two genes. The contents of polyphenols in ntwrky33a, ntmy4, and ntwrky33a/ntmyb4 mutants further confirmed that the repression of NtWRKY33a on the biosynthesis of rutin, scopoletin, and total polyphenols depends on the activity of NtMYB4. Moreover, the promotion of NtHCT by NtWRKY33a modulates the distribution of metabolism flux into the synthesis of CGA. Ectopic expression of NtWRKY33a inhibit the expression of NtSAUR14, NtSAUR59, NtSAUR66, NtIAA4, NtIAA17, and NtIAA19 genes, indicating that NtWRKY33a might be involved in the regulation of plant auxin response. Our study revealed new functions of NtWRKY33a in regulating the synthesis of polyphenols, and provided a promising target for manipulating polyphenols contents in tobacco.
Asunto(s)
Nicotiana , Polifenoles , Nicotiana/genética , Nicotiana/metabolismo , Polifenoles/metabolismo , Factores de Transcripción/genética , Factores de Transcripción/metabolismo , Rutina/metabolismo , Ácido Clorogénico/metabolismo , Escopoletina/metabolismo , Regulación de la Expresión Génica de las PlantasRESUMEN
Coumarins and furanocoumarins are plant secondary metabolites with known biological activities. As they are present in low amounts in plants, their heterologous production emerged as a more sustainable and efficient approach to plant extraction. Although coumarins biosynthesis has been positively established, furanocoumarin biosynthesis has been far more challenging. This study aims to evaluate if Escherichia coli could be a suitable host for furanocoumarin biosynthesis. The biosynthetic pathway for coumarins biosynthesis in E. coli was effectively constructed, leading to the production of umbelliferone, esculetin and scopoletin (128.7, 17.6, and 15.7 µM, respectively, from tyrosine). However, it was not possible to complete the pathway with the enzymes that ultimately lead to furanocoumarins production. Prenyltransferase, psoralen synthase, and marmesin synthase did not show any activity when expressed in E. coli. Several strategies were tested to improve the enzymes solubility and activity with no success, including removing potential N-terminal transit peptides and expression of cytochrome P450 reductases, chaperones and/or enzymes to increase dimethylallylpyrophosphate availability. Considering the results herein obtained, E. coli does not seem to be an appropriate host to express these enzymes. However, new alternative microbial enzymes may be a suitable option for reconstituting the furanocoumarins pathway in E. coli. Nevertheless, until further microbial enzymes are identified, Saccharomyces cerevisiae may be considered a preferred host as it has already been proven to successfully express some of these plant enzymes.
Asunto(s)
Furocumarinas , Furocumarinas/química , Escherichia coli/metabolismo , Cumarinas/metabolismo , Escopoletina/metabolismo , Plantas/metabolismoRESUMEN
Cassava storage roots are an important source of food, feed, and material for starch-based industries in many countries. After harvest, rapid post-harvest physiological deterioration (PPD) reduces their palatability and marketability. During the PPD process, vascular streaking occurs through over-accumulation of coumarins, the biosynthesis of which involves the key enzyme p-coumaroyl shikimate/quinate 3'-hydroxylase (C3'H). Repression of MeC3'H expression by RNA interference in transgenic cassava plants caused a significant delay in PPD by decreasing scopoletin and scopolin accumulation in field-harvested storage roots. This study demonstrates that MeC3'H is the key enzyme participating in coumarin biosynthesis during PPD and shows that MeC3'H is a useful target gene for editing to prolong the shelf life of cassava storage roots.
Asunto(s)
Manihot , Manihot/metabolismo , Oxigenasas de Función Mixta/genética , Raíces de Plantas/metabolismo , Plantas Modificadas Genéticamente/genética , Plantas Modificadas Genéticamente/metabolismo , Ácido Quínico/metabolismo , Escopoletina/metabolismoRESUMEN
Leucine-rich repeat receptor-like kinases (LRR-RKs), belonging to the largest subfamily of transmembrane receptor-like kinases in plants, are proposed to be involved in pathogen resistance. However, it is currently unknown whether LRR-RKs regulate Nicotiana attenuata resistance to Alternaria alternata, a notorious fungal pathogen causing tobacco brown disease. During transcriptome analysis, we identified a highly induced receptor kinase (NaLRR-RK4) in N. attenuata leaves after A. alternata inoculation. We speculated that this NaLRR-RK4 might be the resistance gene of tobacco to brown spot disease, and if so, what is its function and mechanism of action? Silencing of NaLRR-RK4 via virus-induced gene silencing (VIGS) lead to plants highly susceptible to A. alternata, and this result was further confirmed by two stable transformation lines (NaLRR-RK4-RNAi lines) generated by RNA interference technology. The susceptible of NaLRR-RK4-RNAi lines to A. alternata was associated with reduced levels of phytoalexin scopoletin and its key synthesis gene NaF6'H1. Further transcriptome analysis of leaves of WT and NaLRR-RK4-RNAi line after A. alternata inoculation revealed that NaLRR-RK4 regulated NaERF109 and NaDEF19. Silencing NaERF109 or NaDEF19 by VIGS lead to plants more susceptible to A.alternata, demonstrating their role in pathogen resistance. Interestingly, A.alternata-induced expression of NaF6'H1 and NaDEF19 were dramatically reduced in NaERF109-silenced VIGS plants. Taken all together, we identified LRR-RK4 as the first Leucine-rich repeat receptor-like kinases involved in A.alternata resistance in tobacco species, by regulating NaERF109, and subsequently NaDEF19 and NaF6'H1.
Asunto(s)
Nicotiana , Escopoletina , Alternaria/fisiología , Leucina/metabolismo , Plantas , Escopoletina/metabolismo , Sesquiterpenos , Nicotiana/metabolismo , FitoalexinasRESUMEN
Scopoletin is a typical example of coumarins, which can be produced in plants. Scopoletin acts as a precursor for pharmaceutical and health care products, and also possesses promising biological properties, including antibacterial, anti-tubercular, anti-hypertensive, anti-inflammatory, anti-diabetic, and anti-hyperuricemic activity. Despite the potential benefits, the production of scopoletin using traditional extraction processes from plants is unsatisfactory. In recent years, synthetic biology has developed rapidly and enabled the effective construction of microbial cell factories for production of high value-added chemicals. Herein, this review summarizes the progress of scopoletin biosynthesis in artificial microbial cell factories. The two main pathways of scopoletin biosynthesis are summarized firstly. Then, synthetic microbial cell factories are reviewed as an attractive improvement strategy for biosynthesis. Emerging techniques in synthetic biology and metabolic engineering are introduced as innovative tools for the efficient synthesis of scopoletin. This review showcases the potential of biosynthesis of scopoletin in artificial microbial cell factories.
Asunto(s)
Ingeniería Metabólica , Escopoletina , Ingeniería Metabólica/métodos , Plantas , Escopoletina/metabolismo , Biología SintéticaRESUMEN
Scopoletin, also known as 6-methoxy-7 hydroxycoumarin, is one of the naturally occurring coumarin commonly found in many edible plants and plays an important role in human health. Despite the various potential pharmacological properties, the biosynthesis process, method of extraction, and mechanism of action on this compound have not been documented well. In this current review, the biosynthesis pathway, distribution of scopoletin in the plant kingdom, and extraction techniques are elaborated. The in vitro, in vivo, and in silico pharmacological studies are also discussed on antioxidant, antimicrobial, anticancer, anti-inflammation, and neuroprotective aspects of scopoletin. This study may help to understand the benefit of scopoletin containing plants and would be beneficial for the prevention and treatment of diseases.
Asunto(s)
Escopoletina , Humanos , Escopoletina/metabolismo , Escopoletina/farmacologíaRESUMEN
OBJECTIVES: In this study, we investigated whether scopoletin stimulated the secretion of insulin in pancreatic ß cells as well as the underlying mechanism involved in this process. METHODS: We incubated the INS-1 pancreatic ß cells with various concentrations of glucose (1.1, 5.6 or 16.7 mM) in the presence or absence of scopoletin. We then analysed the secretion of insulin in the cells treated with insulin secretion inhibitors or secretagogues. The intracellular influx of calcium induced by scopoletin was also analysed using the Fluo-2 AM dye. KEY FINDINGS: We found that scopoletin (1-20 µM) markedly induced the secretion of insulin in a glucose concentration-dependent manner compared with the control. At depolarizing concentrations of potassium chloride (KCl), scopoletin markedly enhanced the insulin secretion compared with the cells which were treated only with KCl. Moreover, the treatment with diazoxide-opening K+ATP channel and verapamil blocking Ca2+ channel significantly decreased the scopoletin-induced increase in insulin secretion. After the pre-treatment of cells with a Ca2+ fluorescent dye, treatment with 20 µM scopoletin resulted in a significant increase in the influx of intracellular Ca2+, exhibiting fluorescence changes in various spectra. CONCLUSIONS: Scopoletin stimulates the secretion of insulin via a K+ATP channel-dependent pathway in the INS-1 pancreatic ß cells.
Asunto(s)
Células Secretoras de Insulina , Islotes Pancreáticos , Adenosina Trifosfato/metabolismo , Calcio/metabolismo , Glucosa/metabolismo , Insulina/metabolismo , Insulina/farmacología , Células Secretoras de Insulina/metabolismo , Canales KATP , Escopoletina/metabolismo , Escopoletina/farmacologíaRESUMEN
KEY MESSAGE: Overexpression of genes involved in coumarin production and secretion can mitigate mycorrhizal incompatibility in nonhost Arabidopsis plants. The coumarin scopoletin, in particular, stimulates pre-penetration development and metabolism in mycorrhizal fungi. Although most plants can benefit from mutualistic associations with arbuscular mycorrhizal (AM) fungi, nonhost plant species such as the model Arabidopsis thaliana have acquired incompatibility. The transcriptional response of Arabidopsis to colonization by host-supported AM fungi switches from initial AM recognition to defense activation and plant growth antagonism. However, detailed functional information on incompatibility in nonhost-AM fungus interactions is largely missing. We studied interactions between host-sustained AM fungal networks of Rhizophagus irregularis and 18 Arabidopsis genotypes affected in nonhost penetration resistance, coumarin production and secretion, and defense (salicylic acid, jasmonic acid, and ethylene) and growth hormones (auxin, brassinosteroid, cytokinin, and gibberellin). We demonstrated that root-secreted coumarins can mitigate incompatibility by stimulating fungal metabolism and promoting initial steps of AM colonization. Moreover, we provide evidence that major molecular defenses in Arabidopsis do not operate as primary mechanisms of AM incompatibility nor of growth antagonism. Our study reveals that, although incompatible, nonhost plants can harbor hidden tools that promote initial steps of AM colonization. Moreover, it uncovered the coumarin scopoletin as a novel signal in the pre-penetration dialogue, with possible implications for the chemical communication in plant-mycorrhizal fungi associations.
Asunto(s)
Arabidopsis/microbiología , Hongos/crecimiento & desarrollo , Micorrizas/crecimiento & desarrollo , Escopoletina/metabolismo , Arabidopsis/genética , Arabidopsis/metabolismo , Genes de Plantas , Genotipo , Interacciones Microbiota-Huesped/genética , Raíces de Plantas/metabolismo , Raíces de Plantas/microbiología , Transducción de SeñalRESUMEN
AIMS: Scopoletin is a natural anticarcinogenic and antiviral coumarin component. Many studies have proved its anti-cancer effect, and after the preliminary screening of this study, Scopoletin had the best inhibitory effect on Non-small cell lung cancer (NSCLC). But its mechanism for treating NSCLC is still unclear. Therefore, network pharmacology and molecular docking technology were used to explore the potential anti-NSCLC targets and pathways of Scopoletin. The results were verified in vitro. MAIN METHODS: First, Scopoletin was isolated from Fennel and screened to conduct cell proliferation assay on Human lung cancer cell line A549, Human colon cancer cell line HCT-116 and Human hepatoma cell line HepG2 respectively, through the MTT test. Then, the key targets and related pathways were screened through Protein-protein Interaction (PPI) network and "component-target-pathway" (C-TP) network constructed by network pharmacology. And the key targets were selected to dock with Scopoletin via molecular docking. A549 and Human normal lung epithelial cell BEAS-2B were used to verify the results, finally. KEY FINDINGS: Through MTT, A549 was chosen as the test cancer cell. From network pharmacology, 16 targets, 27 signaling pathways and 16 GO items were obtained (P < 0.05). The results of PPI network and molecular docking showed that EGFR, BRAF and AKT1 were the key targets of Scopoletin against NSCLC, which were consistent with the western-blot results. SIGNIFICANCE: Through network pharmacology, molecular docking and experiments in vitro, Scopoletin was verified to against NSCLC through RAS-RAF-MEK-ERK pathway and PI3K/AKT pathway.
Asunto(s)
Carcinoma de Pulmón de Células no Pequeñas/metabolismo , Escopoletina/farmacología , Células A549 , Carcinoma de Pulmón de Células no Pequeñas/tratamiento farmacológico , Línea Celular Tumoral , China , Medicamentos Herbarios Chinos/farmacología , Células HCT116 , Células Hep G2 , Humanos , Medicina Tradicional China/métodos , Simulación del Acoplamiento Molecular/métodos , Mapas de Interacción de Proteínas/efectos de los fármacos , Escopoletina/metabolismo , Transducción de Señal/efectos de los fármacosRESUMEN
Catechol-O-methyltransferase (COMT) methylates both endogenous and exogenous catechol compounds to inactive and safe metabolites. We first optimised conditions for a convenient and sensitive continuous fluorescence-based 6-O-methylation assay of esculetin, which we used for investigating the COMT activity in human, mouse, rat, dog, rabbit, and sheep liver cytosols and microsomes and in ten different rat tissues. Furthermore, we compared the inhibition potencies and mechanisms of two clinically used COMT inhibitors, entacapone and tolcapone, in these species. In most tissues, the COMT activity was at least three times higher in cytosol than in microsomes. In the rat, the highest COMT activity was found in the liver, followed by kidney, ileum, thymus, spleen, lung, pancreas, heart, brain, and finally, skeletal muscle. Entacapone and tolcapone were characterised as highly potent mixed type tight-binding inhibitors. The competitive inhibition type dominated over the uncompetitive inhibition with entacapone, whereas uncompetitive inhibition dominated with tolcapone. Rats, dogs, pigs, and sheep are high COMT activity species, in contrast to humans, mice, and rabbits; COMT activity is highest in the liver. Both entacapone and tolcapone are potent COMT inhibitors, but their inhibition mechanisms differ.