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1.
Artigo em Inglês | MEDLINE | ID: mdl-31642365

RESUMO

This work evaluated the potential of Catharanthus roseus in phytoremediation of As, Ba, Cd, Cu, Cr, Ni, Pb, Se and Zn in sewage sludge-based substrates. C. roseus was cultivated for 108 days in a treatment containing sewage sludge:vermiculite (70:30%) and in the control with 100% commercial substrate. The plants cultivated in sludge showed approximately four times greater height, number of leaves and stem diameter, as well as 89% higher fresh mass than those of the control. The highest concentrations of the metals were obtained in the roots of plants grown in the sludge, and ranged from 2.04 (Cd) to 1121 mg kg-1 (Zn). Cu, Cd and Zn had a higher bioconcentration factor than 1 in both treatments. On the other hand, the translocation factor value in the control was greater than 1 for Ba, Cd, Cu, Ni, Se and Zn. The results showed that C. roseus was efficient in the phytoremediation of the evaluated metals, in which the translocation process was progressive in the treatment that presented higher metal content.


Assuntos
Catharanthus/metabolismo , Metais Pesados/metabolismo , Eliminação de Resíduos/métodos , Poluentes do Solo/metabolismo , Biodegradação Ambiental , Catharanthus/crescimento & desenvolvimento , Metais Pesados/análise , Folhas de Planta/crescimento & desenvolvimento , Folhas de Planta/metabolismo , Raízes de Plantas/crescimento & desenvolvimento , Raízes de Plantas/metabolismo , Esgotos/química , Poluentes do Solo/análise
2.
Int J Syst Evol Microbiol ; 70(2): 964-970, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-31730033

RESUMO

Strain CR1-09T, an actinomycete isolated from the root of Catharanthus roseus, was taxonomically studied based upon polyphasic approaches. The isolate formed a pair of ovular to circular, smooth-surfaced spores on short sporophores alternately branched from aerial mycelia. It contained meso-diaminopimelic acid in cell wall peptidoglycans. The major menaquinones were MK-9 (H4) and MK-9 (H2). The predominant cellular fatty acids were iso-C16 : 0 and C16 : 0. The polar lipids profile consisted of diphosphatidylglycerol, phosphatidylglycerol, phosphatidylethanolamine, hydroxyl phosphatidylethanolamine, phosphatidylinositol, phosphatidylinositol mannosides, and unidentified ninhydrin positive phosphoglycolipids. Strain CR1-09T showed the highest 16S rRNA gene sequence similarity with Microbispora tritici DSM 104650T (99.5 %). Based on the polyphasic approach, DNA-DNA relatedness and average nucleotide identity (ANI), the strain is considered to represent a novel species of the genus Microbispora, for which the name Microbispora catharanthi is proposed. The type strain is strain CR1-09T (=JCM 30045T=TISTR 2273T).


Assuntos
Actinobacteria/classificação , Catharanthus/microbiologia , Filogenia , Raízes de Plantas/microbiologia , Actinobacteria/isolamento & purificação , Técnicas de Tipagem Bacteriana , Composição de Bases , Parede Celular/química , DNA Bacteriano/genética , Ácido Diaminopimélico/química , Ácidos Graxos/química , Hibridização de Ácido Nucleico , Peptidoglicano/química , Fosfolipídeos/química , RNA Ribossômico 16S/genética , Análise de Sequência de DNA , Tailândia , Vitamina K 2/análogos & derivados , Vitamina K 2/química
3.
Environ Sci Pollut Res Int ; 27(7): 6981-6994, 2020 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-31883077

RESUMO

In this study, a pot experiment was performed to evaluate the effects of foliar spray with sodium nitroprusside (200 µM SNP) and melatonin (100 µM) singly and in combination on tolerance and accumulation of cadmium (Cd) in Catharanthus roseus (L.) G. Don plants exposed to different levels of cadmium (0, 50, 100, and 200 mg Cd kg-1 soil). The results showed that 50 mg kg-1 Cd had no significant effect on the fresh and dry weight of roots and shoots and content of chlorophyll (Chl) a and b, but the higher levels of Cd (100 and 200 mg kg-1) significantly reduced these attributes and induced an increase in the level of leaf electrolyte leakage and disrupted nutrient homeostasis. The activities of catalase (CAT) and peroxidase (POD) in leaves were increased under lower Cd concentrations (50 and 100 mg kg-1) but decreased under 200 mg kg-1 Cd. However, foliar spray with melatonin and/or SNP increased shoot biomass and the content of Chl a and b, augmented activities of POD and CAT, lowered electrolyte leakage (EL), and improved essential cations homeostasis in leaves. Cadmium content in shoots of C. roseus was less than roots and TF (transfer factor) was < 1. Interestingly, foliar spray with SNP and/or melatonin increased Cd accumulation and bioconcentration factor (BCF) in both roots and shoots and elevated the Cd transport from roots to shoot, as TF values increased in these treatments. The co-application of melatonin and SNP further than their separate usage augmented Cd tolerance through increasing activities of antioxidant enzymes and regulating mineral homeostasis in C. roseus. Furthermore, co-treatment of SNP and melatonin increased Cd phytoremediation efficiency in C. roseus through increasing biomass and elevating uptake and translocation of Cd from root to shoot.


Assuntos
Biodegradação Ambiental , Cádmio/toxicidade , Catharanthus , Melatonina/metabolismo , Óxido Nítrico/metabolismo , Poluentes do Solo/toxicidade , Antioxidantes , Raízes de Plantas
4.
Plant Sci ; 290: 110293, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31779893

RESUMO

Class I TGA transcription factors (TFs) are known to participate in plant resistance responses, however, their regulatory functions in the biosynthesis of secondary metabolites were rarely revealed. In this study, a class I TGA TF, TwTGA1, from Tripterygium wilfordii Hook.f. was cloned and characterized. Overexpression of TwTGA1 in T. wilfordii Hook.f. cells increased the production of triptolide and two sesquiterpene pyridine alkaloids, which was further enhanced by methyl jasmonate (MeJA) treatment. RNA interference of TwTGA1 showed no significant effects on the production of these metabolites, indicating the existence of other TGA partner(s) with overlapping functions. Heterologous expression of TwTGA1 in tobacco By-2 cells promoted the biosynthesis of pyridine alkaloids. Under the elicitation of MeJA, the contents of nonpyrrolidine alkaloids further increased but not for nicotine. TwTGA1 could induce the expression of Putrescine N-methyltransferase (PMT) and N-methylputrescine oxidase 1 (MPO1) through binding to their promoters. Finally, transient expression of TwTGA1 in leaves of Catharanthus roseus changed both the profiles of vinca alkaloids (increased contents of serpentine and catharanthine, but decreased that of vinblastine) and the expressions of biosynthesis-related genes. The metabolic and transcriptional data indicated a relationship between jasmonic acid signaling pathway and the functions of TwTGA1.


Assuntos
Regulação da Expressão Gênica de Plantas , Proteínas de Plantas/genética , Fatores de Transcrição/genética , Tripterygium/genética , Alcaloides/biossíntese , Sequência de Aminoácidos , Catharanthus/metabolismo , Diterpenos/metabolismo , Compostos de Epóxi/metabolismo , Fenantrenos/metabolismo , Filogenia , Proteínas de Plantas/química , Proteínas de Plantas/metabolismo , Plantas Geneticamente Modificadas/metabolismo , Metabolismo Secundário , Alinhamento de Sequência , Fatores de Transcrição/química , Fatores de Transcrição/metabolismo , Tripterygium/metabolismo
5.
BMC Complement Altern Med ; 19(1): 338, 2019 Nov 29.
Artigo em Inglês | MEDLINE | ID: mdl-31783835

RESUMO

BACKGROUND: To investigate the potential of Catharanthus roseus leaf aqueous crude extract (CRACE) as a regulator of adipocyte development and function. METHODS: 3T3-L1 adipogenesis model was used to investigate the effect of CRACE on adipogenesis. 3T3-L1 preadipocytes (for adipogenic differentiation) and mature 3T3-L1 adipocytes (for adipocyte function) were treated with non-toxic doses of CRACE. The outcomes were corroborated by intracellular lipid accumulation, expression of pro-and anti-adipogenic effector molecules. To investigate CRACE mediated lipolysis, cAMP accumulation, glycerol release and phosphorylation of key effector molecules were tested in treated mature adipocytes. Finally, the extract was fractionated to identify the active molecule/s in the extract. RESULTS: CRACE significantly reduced adipocyte differentiation by modulating PPARγ expression. At early stage CRACE directly targeted Lipin1 expression and consequently impacted KLF7, subsequently expression of GATA2, CEBPα, SREBP1c were targeted, with PPARγ expression, particularly curtailed. While CRACE significantly reduced several lipogenic genes like FAS and GPD1 in mature adipocytes, concomitantly, it greatly increased lipolysis resulting in decreased lipid accumulation in mature adipocytes. The increase in lipolysis was due to decreased Akt activation, increased cAMP level, and PKA activity. The fractionation of CRACE allowed identification of two fractions with potent anti-adipogenic activity. Both the fractions contained 1α, 25-dihydroxy Vitamin D3 as major component. CONCLUSIONS: 1α, 25-dihydroxy Vitamin D3 containing CRACE can be developed into an effective anti-obesity formulation that decreases adipogenesis and increases lipid catabolism.


Assuntos
Adipócitos/efeitos dos fármacos , Adipogenia/efeitos dos fármacos , Calcitriol/farmacologia , Catharanthus , Lipólise/efeitos dos fármacos , Células 3T3-L1 , Animais , Camundongos , Extratos Vegetais/farmacologia , Folhas de Planta/química
6.
Planta ; 251(1): 13, 2019 Nov 27.
Artigo em Inglês | MEDLINE | ID: mdl-31776675

RESUMO

MAIN CONCLUSION: Phenolic oxidative coupling protein (Hyp-1) isolated from Hypericum perforatum L. was characterized as a defense gene involved in H. perforatum recalcitrance to Agrobacterium tumefaciens-mediated transformation Hypericum perforatum L. is a reservoir of high-value secondary metabolites of increasing interest to researchers and to the pharmaceutical industry. However, improving their production via genetic manipulation is a challenging task, as H. perforatum is recalcitrant to Agrobacterium tumefaciens-mediated transformation. Here, phenolic oxidative coupling protein (Hyp-1), a pathogenesis-related (PR) class 10 family gene, was selected from a subtractive cDNA library from A. tumefaciens-treated H. perforatum suspension cells. The role of Hyp-1 in defense against A. tumefaciens was analyzed in transgenic Nicotiana tabacum and Lactuca sativa overexpressing Hyp-1, and in Catharanthus roseus silenced for its homologous Hyp-1 gene, CrIPR. Results showed that Agrobacterium-mediated expression efficiency greatly decreased in Hyp-1 transgenic plants. However, silencing of CrIPR induced CrPR-5 expression and decreased expression efficiency of Agrobacterium. The expression of core genes involved in several defense pathways was also analyzed in Hyp-1 transgenic tobacco plants. Overexpression of Hyp-1 led to an ample down-regulation of key genes involved in auxin signaling, microRNA-based gene silencing, detoxification of reactive oxygen species, phenylpropanoid pathway and PRs. Moreover, Hyp-1 was detected in the nucleus, plasma membrane and the cytoplasm of epidermal cells by confocal microscopy. Overall, our findings suggest Hyp-1 modulates recalcitrance to A. tumefaciens-mediated transformation in H. perforatum.


Assuntos
Agrobacterium tumefaciens/fisiologia , Catharanthus/metabolismo , Hypericum/metabolismo , Catharanthus/microbiologia , Hypericum/microbiologia , Plantas Geneticamente Modificadas/metabolismo , Plantas Geneticamente Modificadas/microbiologia , Tabaco/metabolismo , Tabaco/microbiologia
7.
Phytochemistry ; 168: 112110, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31494345

RESUMO

Based on the occurrence of indole alkaloids in so-called "chloroform leaf surface extracts", it was previously deduced that these alkaloids are present in the cuticle at the leaf surface of Catharanthus roseus and Vinca minor. As no symplastic markers were found in these extracts this deduction seemed to be sound. However, since chloroform is known to destroy biomembranes very rapidly, these data have to be judged with scepticism. We reanalyzed the alleged apoplastic localization of indole alkaloids by employing slightly acidic aqueous surface extracts and comparing the corresponding alkaloid patterns with those of aqueous total leaf extracts. Whereas in the "chloroform leaf surface extracts" all alkaloids are present in the same manner as in the total leaf extracts, no alkaloids occur in the aqueous leaf surface extracts. These results clearly show that chloroform had rapidly destroyed cell integrity, and the related extracts also contain the alkaloids genuinely accumulated within the protoplasm. The related decompartmentation was verified by the massively enhanced concentration of amino acids in aqueous surface extracts of chloroform treated leaves. Furthermore, the chloroform-induced cell disintegration was vividly visualized by confocal laser scanning microscopical analyses, which clearly displayed a strong decrease in the chlorophyll fluorescence in chloroform treated leaves. These findings unequivocally display that the indole alkaloids are not located in the apoplastic space, but exclusively are present symplastically within the cells of V. minor and C. roseus leaves. Accordingly, we have to presume that also other leaf surface extracts employing organic solvents have to be re-investigated.


Assuntos
Catharanthus/química , Alcaloides Indólicos/análise , Alcaloides Indólicos/isolamento & purificação , Extratos Vegetais/análise , Extratos Vegetais/isolamento & purificação , Folhas de Planta/citologia , Vinca/química , Catharanthus/citologia , Alcaloides Indólicos/química , Extratos Vegetais/química , Folhas de Planta/química , Vinca/citologia
8.
New Phytol ; 224(2): 848-859, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31436868

RESUMO

Catharanthus roseus is a medicinal plant well known for producing bioactive compounds such as vinblastine and vincristine, which are classified as terpenoid indole alkaloids (TIAs). Although the leaves of this plant are the main source of these antitumour drugs, much remains unknown on how TIAs are biosynthesised from a central precursor, strictosidine, to various TIAs in planta. Here, we have succeeded in showing, for the first time in leaf tissue of C. roseus, cell-specific TIAs localisation and accumulation with 10 µm spatial resolution Imaging mass spectrometry (Imaging MS) and live single-cell mass spectrometry (single-cell MS). These metabolomic studies revealed that most TIA precursors (iridoids) are localised in the epidermal cells, but major TIAs including serpentine and vindoline are localised instead in idioblast cells. Interestingly, the central TIA intermediate strictosidine also accumulates in both epidermal and idioblast cells of C. roseus. Moreover, we also found that vindoline accumulation increases in laticifer cells as the leaf expands. These discoveries highlight the complexity of intercellular localisation in plant specialised metabolism.


Assuntos
Catharanthus/citologia , Catharanthus/metabolismo , Metabolômica , Folhas de Planta/citologia , Alcaloides de Triptamina e Secologanina/metabolismo , Técnicas de Cultura de Células , Análise de Componente Principal
9.
Metab Eng ; 55: 76-84, 2019 09.
Artigo em Inglês | MEDLINE | ID: mdl-31226348

RESUMO

Monoterpene indole alkaloids (MIAs) from plants encompass a broad class of structurally complex and medicinally valuable natural products. MIAs are biologically derived from the universal precursor strictosidine. Although the strictosidine biosynthetic pathway has been identified and reconstituted, extensive work is required to optimize production of strictosidine and its precursors in yeast. In this study, we engineered a fully integrated and plasmid-free yeast strain with enhanced production of the monoterpene precursor geraniol. The geraniol biosynthetic pathway was targeted to the mitochondria to protect the GPP pool from consumption by the cytosolic ergosterol pathway. The mitochondrial geraniol producer showed a 6-fold increase in geraniol production compared to cytosolic producing strains. We further engineered the monoterpene-producing strain to synthesize the next intermediates in the strictosidine pathway: 8-hydroxygeraniol and nepetalactol. Integration of geraniol hydroxylase (G8H) from Catharanthus roseus led to essentially quantitative conversion of geraniol to 8-hydroxygeraniol at a titer of 227 mg/L in a fed-batch fermentation. Further introduction of geraniol oxidoreductase (GOR) and iridoid synthase (ISY) from C. roseus and tuning of the relative expression levels resulted in the first de novo nepetalactol production. The strategies developed in this work can facilitate future strain engineering for yeast production of later intermediates in the strictosidine biosynthetic pathway.


Assuntos
Engenharia Metabólica , Microrganismos Geneticamente Modificados , Mitocôndrias , Monoterpenos/metabolismo , Saccharomyces cerevisiae , Alcaloides de Vinca/biossíntese , Catharanthus/enzimologia , Catharanthus/genética , Microrganismos Geneticamente Modificados/genética , Microrganismos Geneticamente Modificados/metabolismo , Mitocôndrias/genética , Mitocôndrias/metabolismo , Proteínas Mitocondriais/genética , Proteínas Mitocondriais/metabolismo , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismo
10.
Artif Cells Nanomed Biotechnol ; 47(1): 1938-1946, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31099261

RESUMO

Bionanotechnology has pivotal role in the development of a novel therapy, applications of gold nanoparticles (AuNPs) in the treatment of cancer. In this study, we found that therapeutics, pharmaceutics and diagnostic effectiveness of photosynthesized Catharanthus roseus (CR) AuNPs induces mitochondrial-mediated apoptotic signalling pathways via reactive oxygen species (ROS) induced cytotoxicity in cervical cancer cell line (HeLa) by in vitro model. The present examinations were for the most part centred around the gold chloride and photosynthesis AuNPs from the fluid leaf concentrate of CR and their harmful impacts on HeLa cell lines. The synthesized AuNPs were characterized using numerous biophysical analyses such as UV-vis, DLS, EDX, HR-TEM, SAED, FTIR and AFM. The synthesized AuNPs in the particle size range of 25-35 nm was confirmed by HR-TEM. The element gold and the crystalline nature of AuNPs were finalized using EDX, respectively. Anticancer potential of CR-AuNPs was studied using HeLa cells and the cytotoxic mechanism has been evaluated using MTT, mitochondrial-mediated apoptotic pathway through AO/EtBr staining assay, pro-apoptotic (Bax), anti-apoptotic (Bcl-2 and Bid) protein expression western blotting analysis and caspases activity using ELISA analysis. In in vitro study, the IC50 of HeLa cells was found to be 5 µg/ml confirmed using MTT assay. The present data revealed that drug delivery vehicles developed on CR-AuNPs nanocomplexes might include extensive purpose in human cancer diagnosis and treatment.


Assuntos
Caspase 3/metabolismo , Caspase 9/metabolismo , Catharanthus/química , Ouro/química , Ouro/farmacologia , Nanopartículas Metálicas/química , Nanotecnologia , Antineoplásicos/síntese química , Antineoplásicos/química , Antineoplásicos/farmacologia , Feminino , Química Verde , Células HeLa , Humanos , Espaço Intracelular/efeitos dos fármacos , Espaço Intracelular/metabolismo , Fotossíntese , Extratos Vegetais/química , Folhas de Planta/química , Espécies Reativas de Oxigênio/metabolismo , Neoplasias do Colo do Útero/patologia
11.
Mol Biol Rep ; 46(3): 3265-3273, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-30945069

RESUMO

Catharanthus roseus (L.) G. Don (C. roseus) is a well-known medicinal plant for its source of alkaloids solely found in the leaves. Other parts including the root are usually discarded after the alkaloid extraction. This study sought to investigate phytochemical profiles, antioxidant, antimicrobial and cytotoxic properties of the C. roseus root extract (RE) and its two sub-fractions including saponin-enriched (SE) and aqueous (AQ) fractions. The results showed that the RE was a rich source of saponins (1744.44 mg ESE/g) and phenolics (51.27 mg GAE/g), which comprised of gallic acid (25.74 mg/g), apigenin (1.45 mg/g) and kaempferol (1.58 mg/g). The SE fraction was enriched with 31% of saponins and 63% of phenolics higher than those of the RE; whereas the concentrations of saponins and phenolics of the AQ fraction were lower than those of the RE by 40% and 74%, respectively. The content of gallic acid in the SE fraction was 1.4-fold and 1.5-fold higher than those of the RE or AQ fraction, respectively. The SE fraction demonstrated potent antioxidant capacity, which was significantly higher than the RE or AQ fraction, and also exhibited strong anti-proliferative activity against 11 cancer cell lines including A2780 (ovarian), H460 (lung), A431 (skin), MIA PaCa-2 (pancreas), Du145 (prostate), HT29 (colon), MCF-7 (breast), BE2-C (neuroblastoma), SJ-G2, U87 and SMA (glioblastoma) with low GI50 values (≤ 2.00 µg/mL). The SE fraction was also shown to effectively inhibit the growth of both bacteria (Escherichia coli, Enterobacter aerogenes and Staphylococccus lugdunensis) and fungi (Candida albicans and Aspergillus niger). These findings warrant further investigation to isolate major compounds from the SE fraction and further test their antioxidant, anticancer and antimicrobial activities.


Assuntos
Catharanthus/química , Catharanthus/metabolismo , Extratos Vegetais/farmacologia , Alcaloides/farmacologia , Antibacterianos/farmacologia , Anti-Infecciosos/farmacologia , Antioxidantes/metabolismo , Linhagem Celular Tumoral/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Flavonoides/farmacologia , Humanos , Fenóis , Compostos Fitoquímicos/metabolismo , Extratos Vegetais/metabolismo , Raízes de Plantas/metabolismo , Plantas Medicinais/metabolismo , Saponinas
12.
Nanotechnology ; 30(27): 275102, 2019 Jul 05.
Artigo em Inglês | MEDLINE | ID: mdl-30901766

RESUMO

Carbon-based nanomaterials (CBNs) were previously described as regulators of plant cell division. Here, we demonstrated the ability of multi-walled carbon nanotubes (MWCNT) and graphene to enhance biomass production in callus culture of the medicinal plant Catharanthus roseus cultivated in dark conditions. Furthermore, both tested CBNs were able to stimulate biosynthesis of total produced alkaloids in CBN-exposed callus culture of Catharanthus. In one case, total alkaloids in CBN-exposed Catharanthus were double that of unexposed Catharanthus. Analysis of metabolites by HPLC revealed that production of the pharmaceutically active alkaloids vinblastine and vincristine was dramatically enhanced in callus exposed to MWCNT or graphene in both dark and light conditions of callus cultivation. In vitro assays (MTT, flow cytometry) demonstrated that total alkaloid extracts derived from Catharanthus callus treated with CBNs significantly reduced cell proliferation of breast cancer (MCF-7) and lung cancer (A549) cell lines compared to the application of extracts derived from untreated Catharanthus callus.


Assuntos
Alcaloides/biossíntese , Alcaloides/farmacologia , Catharanthus/química , Catharanthus/crescimento & desenvolvimento , Nanotubos de Carbono/química , Células A549 , Catharanthus/efeitos dos fármacos , Técnicas de Cultura de Células , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Grafite/farmacologia , Humanos , Células MCF-7 , Necrose , Extratos Vegetais/farmacologia , Vimblastina/farmacologia
13.
Biomed Pharmacother ; 112: 108638, 2019 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-30784928

RESUMO

Vindoline, an indole alkaloid present in the leaves of Catharanthus roseus plant, has been recently reported to have insulotropic effects. This present study evaluated the possible hepatoprotective effects of vindoline in a type 2 diabetes mellitus rat model. Diabetes mellitus was induced by exposing rats to 10% fructose water for two weeks followed by a single intraperitoneal injection of 40 mg/kg body weight of streptozotocin (STZ). Rats were randomly divided into six groups (n = 8) and treated daily for 6 weeks with the vehicle via oral gavage, vindoline (20 mg/kg) or glibenclamide (5 mg/kg). Weekly fasting blood glucose (FBG) levels and body weight were measured and recorded. Administration of vindoline significantly (p < 0.05) reduced FBG by 15% when compared to the diabetic controls. Vindoline significantly (p < 0.05) decreased diabetes-induced hepatic injury shown by decreased levels of serum alanine transferase (ALT) (-42%), aspartate aminotransferase (AST) (-42%) and alkaline phosphatase (-62%) compared to the diabetic controls. The oxygen radical absorbance capacity and the activities of superoxide dismutase (SOD) and catalase (CAT) were also improved following treatment with vindoline. The results also showed decreased levels of pro-inflammatory cytokines such as TNF-ɑ by (-41%) and IL-6 (-28%) which may have also contributed to the reduction of serum triglycerides (-65%) in the diabetic group treated with vindoline. Histopathological findings showed improvement of both the hepatic and pancreatic tissues following vindoline treatment. Overall, these findings suggest that vindoline may protect the diabetic hepatic tissue from injury via antioxidant, anti-inflammatory and anti-hypertriglyceredemia mechanisms thereby retarding the development of diabetic complications.


Assuntos
Diabetes Mellitus Experimental/complicações , Diabetes Mellitus Tipo 2/complicações , Hepatite/prevenção & controle , Hipertrigliceridemia/prevenção & controle , Hipoglicemiantes/uso terapêutico , Estresse Oxidativo/efeitos dos fármacos , Vimblastina/análogos & derivados , Animais , Glicemia/análise , Catharanthus/química , Citocinas/metabolismo , Diabetes Mellitus Experimental/metabolismo , Diabetes Mellitus Tipo 2/metabolismo , Glibureto/uso terapêutico , Hepatite/imunologia , Hepatite/metabolismo , Hepatite/patologia , Insulina/sangue , Testes de Função Hepática , Masculino , Pâncreas/efeitos dos fármacos , Pâncreas/patologia , Ratos Wistar , Vimblastina/uso terapêutico
14.
BMC Microbiol ; 19(1): 22, 2019 01 21.
Artigo em Inglês | MEDLINE | ID: mdl-30665368

RESUMO

BACKGROUND: The present study involves diversity and bioactivity of the endophytic fungal community from Catharanthus roseus inhabiting the coastal region. This study has been conducted hypothesizing that the microbial communities in the coastal regions would tolerate a range of abiotic stress such as salinity, humidity, temperature and soil composition, and it may produce new metabolites, which may possess bioactive property. Therefore in the current study, the cytotoxicity and free radical scavenging potential of the fungal organic extracts have been investigated. Moreover, the apoptotic and the antioxidant potential of the fungus that exhibited the best activity in preliminary screening has also been demonstrated. RESULTS: Twenty endophytic fungal isolates were obtained from different parts of the plant, and identified using internal transcribed spacer region analysis. Based on the colonization frequency, the dominant genera were found to be Colletotrichum, Alternaria and Chaetomium with colonization frequency % of 8.66, 7.00 and 6.33, respectively. It was observed that the species diversity and richness was the highest in bark followed by leaf and stem regions of the plant. On screening the fungal ethyl acetate extracts for cytotoxicity against the HeLa cells, the Chaetomium nigricolor extract exhibited potent cytotoxic activity of 92.20% at 100 µg mL- 1 concentration. Comparison between the different organic extracts (ethyl acetate, chloroform, dichloromethane and hexane) of Chaetomium nigricolor mycelial and culture filtrate, it was observed that the mycelial as well the culture filtrate ethyl acetate extracts and the culture filtrate hexane extract showed significant cytotoxic potential against the HeLa and MCF-7 cells, respectively. The apoptotic- and mitochondrial membrane depolarisation-induction potential of the Chaetomium nigricolor ethyl acetate extract has also been demonstrated in this study. Further the screening of antioxidant potential of the ethyl acetate fungal extracts using DPPH scavenging assay showed that Chaetomium nigricolor extract exhibited potential activity with a significant EC50 value of 22 µg mL- 1. The ethyl acetate extract of Chaetomium nigricolor also exhibited superoxide radical scavenging potential. CONCLUSION: These results indicated that diverse endophytic fungal population inhabits Catharanthus roseus. One of the fungal isolate Chaetomium nigricolor exhibited significant cytotoxic, apoptotic and antioxidant potential.


Assuntos
Catharanthus/microbiologia , Endófitos/química , Endófitos/classificação , Fungos/química , Fungos/classificação , Apoptose , Endófitos/isolamento & purificação , Depuradores de Radicais Livres/química , Fungos/isolamento & purificação , Células HEK293 , Células HeLa , Humanos , Índia , Células MCF-7 , Potencial da Membrana Mitocondrial , Consórcios Microbianos , Folhas de Planta/microbiologia , Caules de Planta/microbiologia
15.
Phytochemistry ; 159: 119-126, 2019 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-30611871

RESUMO

The medicinal value of the monoterpenoid indole alkaloids (MIAs) such as 3',4'-anhydrovinblastine, as well as their chemical complexity have stimulated extensive efforts to understand the biochemical and molecular pathways involved in their biosynthesis in plants such as Catharanthus roseus, Rawvolfia serpentina and others. Ethyl methane sulphonate (EMS) mutagenesis has been used successfully together with simple MIA thin layer chromatography screening to identify C. roseus mutants with altered MIA profiles. This study describes the isolation of very low iridoid and MIA containing C. roseus mutant (M2-1582) that accumulates MIAs when the plant is provided with secologanin by feeding mutant roots or by grafting the mutant scion onto wild type roots. The observed low iridoid and MIA content was correlated with lowered expression of BIS1/BIS2 transcription factors and several genes involved in secologanin biosynthesis that are expressed in internal phloem parenchyma cells of leaves. When exogenous secologanin was applied to the roots of the mutant plant, secologanin levels rose more than 13-fold, while two major MIAs catharanthine and vindoline rose more than 8- and 4- fold, respectively. Grafting the mutant on WT stocks led to 27-, 11- and 27-fold increases in secologanin, catharanthine and vindoline, respectively in leaves of the scion one week after graft initiation. Other minor MIAs (serpentine, anhydrovinblastine, vindolidine, deacetylvindoline, tabersonine and 16-methoxytabersonine) that were not detected in the mutant, became detectable in leaves of the scion. These results provide strong evidence for a secologanin transport mechanism that mobilizes this iridoid between different plant organs in C. roseus and that secologanin transport to the mutant across the graft union permits the formation of MIAs in leaves of the mutant.


Assuntos
Catharanthus/metabolismo , Alcaloides Indólicos/metabolismo , Glucosídeos Iridoides/metabolismo , Monoterpenos/metabolismo , Mutação , Folhas de Planta/metabolismo , Raízes de Plantas/metabolismo , Brotos de Planta/metabolismo , Fatores de Transcrição/metabolismo , Transporte Biológico , Catharanthus/genética , Cromatografia em Camada Delgada , Regulação para Baixo
16.
Plant J ; 97(2): 257-266, 2019 01.
Artigo em Inglês | MEDLINE | ID: mdl-30256480

RESUMO

The important anticancer drugs, vinblastine, vincristine and analogs, are composed of the monoterpenoid indole alkaloids (MIAs), catharanthine and vindoline, found uniquely in the medicinal plant, Catharanthus roseus. While 26 genes involved in the assembly of these two MIAs are known, two key reactions have eluded characterization to complete the documentation of the vinblastine pathway in this plant species. The assembly of these dimeric MIAs requires O-acetylstemmadenine oxidase (ASO) and a dual function geissoschizine synthase (GS) that reduces cathenamine to form geissoschizine, and that also reduces the ASO product to form a common intermediate for subsequent conversion by four separate hydrolases to catharanthine, tabersonine or vincadifformine, respectively. The in planta role of ASO is supported by identifying a single amino acid-substituted ASO mutant with very low enzyme activity and by virus-induced gene silencing of ASO to produce plants that accumulate O-acetylstemmadenine rather than catharanthine and vindoline found in wild-type (WT) plants. The in planta role of GS is supported by showing that a low GS-expressing mutant accumulating lower levels of catharanthine and vindoline also displays significantly lower tabersonine-forming activity in coupled enzyme assays than in the WT background. Gene expression analyses demonstrate that both ASO and GS are highly enriched in the leaf epidermis where the pathways for catharanthine and tabersonine biosynthesis are expressed. The full elucidation of this canonical pathway enables synthetic biology approaches for manufacturing a broad range of MIAs, including these dimers used in cancer treatment.


Assuntos
Antineoplásicos Fitogênicos/metabolismo , Catharanthus/genética , Proteínas de Plantas/metabolismo , Alcaloides de Triptamina e Secologanina/metabolismo , Vimblastina/metabolismo , Vincristina/metabolismo , Carbolinas/metabolismo , Catharanthus/metabolismo , Alcaloides Indólicos/metabolismo , Engenharia Metabólica , Oxirredutases/genética , Oxirredutases/metabolismo , Proteínas de Plantas/genética , Plantas Medicinais , Biologia Sintética , Vimblastina/análogos & derivados , Alcaloides de Vinca/metabolismo
17.
Syst Appl Microbiol ; 42(2): 117-127, 2019 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-30455068

RESUMO

Bacteria of the genus 'Candidatus Phytoplasma' are uncultivated intracellular plant pathogens transmitted by phloem-feeding insects. They have small genomes lacking genes for essential metabolites, which they acquire from either plant or insect hosts. Nonetheless, some phytoplasmas, such as 'Ca. P. solani', have broad plant host range and are transmitted by several polyphagous insect species. To understand better how these obligate symbionts can colonize such a wide range of hosts, the genome of 'Ca. P. solani' strain SA-1 was sequenced from infected periwinkle via a metagenomics approach. The de novo assembly generated a draft genome with 19 contigs totalling 821,322bp, which corresponded to more than 80% of the estimated genome size. Further completion of the genome was challenging due to the high occurrence of repetitive sequences. The majority of repeats consisted of gene arrangements characteristic of phytoplasma potential mobile units (PMUs). These regions showed variation in gene orders intermixed with genes of unknown functions and lack of similarity to other phytoplasma genes, suggesting that they were prone to rearrangements and acquisition of new sequences via recombination. The availability of this high-quality draft genome also provided a foundation for genome-scale genotypic analysis (e.g., average nucleotide identity and average amino acid identity) and molecular phylogenetic analysis. Phylogenetic analyses provided evidence of horizontal transfer for PMU-like elements from various phytoplasmas, including distantly related ones. The 'Ca. P. solani' SA-1 genome also contained putative secreted protein/effector genes, including a homologue of SAP11, found in many other phytoplasma species.


Assuntos
Ordem dos Genes , Genoma Bacteriano , Phytoplasma/genética , Catharanthus/microbiologia , DNA Bacteriano , Metagenômica , Filogenia
18.
Chembiochem ; 20(1): 83-87, 2019 01 02.
Artigo em Inglês | MEDLINE | ID: mdl-30300974

RESUMO

Genome mining is a routine technique in microbes for discovering biosynthetic pathways. In plants, however, genomic information is not commonly used to identify novel biosynthesis genes. Here, we present the genome of the medicinal plant and oxindole monoterpene indole alkaloid (MIA) producer Gelsemium sempervirens (Gelsemiaceae). A gene cluster from Catharanthus roseus, which is utilized at least six enzymatic steps downstream from the last common intermediate shared between the two plant alkaloid types, is found in G. sempervirens, although the corresponding enzymes act on entirely different substrates. This study provides insights into the common genomic context of MIA pathways and is an important milestone in the further elucidation of the Gelsemium oxindole alkaloid pathway.


Assuntos
Gelsemium/genética , Genes de Plantas , Alcaloides Indólicos/metabolismo , Monoterpenos/metabolismo , Família Multigênica , Catharanthus/genética , Estudos de Associação Genética , Genoma , Raízes de Plantas/genética
19.
Molecules ; 23(12)2018 Dec 11.
Artigo em Inglês | MEDLINE | ID: mdl-30544939

RESUMO

This study reported the inducing effect of Aspergillus flavus fungal elicitor on biosynthesis of terpenoid indole alkaloids (TIAs) in Catharanthus roseus cambial meristematic cells (CMCs) and its inducing mechanism. According to the results determined by HPLC and HPLC-MS/MS, the optimal condition of the A. flavus elicitor was as follows: after suspension culture of C. roseus CMCs for 6 day, 25 mg/L A. flavus mycelium elicitor were added, and the CMC suspensions were further cultured for another 48 h. In this condition, the contents of vindoline, catharanthine, and ajmaline were 1.45-, 3.29-, and 2.14-times as high as those of the control group, respectively. Transcriptome analysis showed that D4H, G10H, GES, IRS, LAMT, SGD, STR, TDC, and ORCA3 were involved in the regulation of this induction process. The results of qRT-PCR indicated that the increasing accumulations of vindoline, catharanthine, and ajmaline in C. roseus CMCs were correlated with the increasing expression of the above genes. Therefore, A. flavus fungal elicitor could enhance the TIA production of C. roseus CMCs, which might be used as an alternative biotechnological resource for obtaining bioactive alkaloids.


Assuntos
Aspergillus flavus/química , Catharanthus/metabolismo , Alcaloides de Triptamina e Secologanina/metabolismo , Catharanthus/química , Catharanthus/citologia , Catharanthus/genética , Cromatografia Líquida de Alta Pressão , Regulação da Expressão Gênica de Plantas , Meristema/citologia , Meristema/metabolismo , Células Vegetais/efeitos dos fármacos , Células Vegetais/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Alcaloides de Triptamina e Secologanina/análise , Espectrometria de Massas em Tandem
20.
ACS Chem Biol ; 13(12): 3343-3353, 2018 12 21.
Artigo em Inglês | MEDLINE | ID: mdl-30484626

RESUMO

Aromatic l-amino acid decarboxylases (AAADs) are a phylogenetically diverse group of enzymes responsible for the decarboxylation of aromatic amino acid substrates into their corresponding aromatic arylalkylamines. AAADs have been extensively studied in mammals and plants as they catalyze the first step in the production of neurotransmitters and bioactive phytochemicals, respectively. Unlike mammals and plants, the hallucinogenic psilocybin mushroom Psilocybe cubensis reportedly employs an unrelated phosphatidylserine-decarboxylase-like enzyme to catalyze l-tryptophan decarboxylation, the first step in psilocybin biosynthesis. To explore the origin of this chemistry in psilocybin mushroom, we generated the first de novo transcriptomes of P. cubensis and investigated several putative l-tryptophan-decarboxylase-like enzymes. We report the biochemical characterization of a noncanonical AAAD from P. cubensis ( PcncAAAD) that exhibits substrate permissiveness toward l-phenylalanine, l-tyrosine, and l-tryptophan, as well as chloro-tryptophan derivatives. The crystal structure of PcncAAAD revealed the presence of a unique C-terminal appendage domain featuring a novel double-ß-barrel fold. This domain is required for PcncAAAD activity and regulates catalytic rate and thermal stability through calcium binding. PcncAAAD likely plays a role in psilocybin production in P. cubensis and offers a new tool for metabolic engineering of aromatic-amino-acid-derived natural products.


Assuntos
Descarboxilases de Aminoácido-L-Aromático/química , Descarboxilases de Aminoácido-L-Aromático/metabolismo , Cálcio/metabolismo , Psilocybe/enzimologia , Psilocibina/biossíntese , Aminoácidos Aromáticos/metabolismo , Descarboxilases de Aminoácido-L-Aromático/genética , Domínio Catalítico , Catharanthus/enzimologia , Proteínas Fúngicas/química , Proteínas Fúngicas/genética , Proteínas Fúngicas/metabolismo , Cinética , Simulação de Acoplamento Molecular , Papaver/enzimologia , Filogenia , Ligação Proteica , Domínios Proteicos , Estrutura Terciária de Proteína , Psilocybe/genética , Especificidade por Substrato , Transcriptoma , Leveduras/genética
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