methyl-jasmonate and vindoline

Long-term stable cell growth and production of vindoline, catharanthine, and ajmalicine of cambial meristematic cells (CMCs) from Catharanthus roseus were observed after 2 years of culture. C. roseus CMCs were treated with β-cyclodextrin (β-CD) and methyl jasmonate (MeJA) individually or in combination and were cultured both in conventional Erlenmeyer flasks (100, 250, and 500 mL) and in a 5-L stirred hybrid airlift bioreactor. CMCs of C. roseus cultured in the bioreactor showed higher yields of vindoline, catharanthine, and ajmalicine than those cultured in flasks. CMCs of C. roseus cultured in the bioreactor and treated with 10 mM β-CD and 150 μM MeJA gave the highest yields of vindoline (7.45 mg/L), catharanthine (1.76 mg/L), and ajmalicine (58.98 mg/L), concentrations that were 799, 654, and 426 % higher, respectively, than yields of CMCs cultured in 100-mL flasks without elicitors. Quantitative reverse transcription (RT)-PCR showed that β-CD and MeJA upregulated transcription levels of genes related to the biosynthesis of terpenoid indole alkaloids (TIAs). This is the first study to report that β-CD induced the generation of NO, which plays an important role in mediating the production of TIAs in C. roseus CMCs. These results suggest that β-CD and MeJA can enhance the production of TIAs in CMCs of C. roseus, and thus, CMCs of C. roseus have significant potential to be an industrial platform for production of bioactive alkaloids.

Topics: Acetates; beta-Cyclodextrins; Catharanthus; Cells, Cultured; Cyclopentanes; Oxylipins; Plant Cells; Secologanin Tryptamine Alkaloids; Vinblastine; Vinca Alkaloids

Vincristine and vinblastine were found by Liquid Chromatography-Mass Spectrometry (LC-MS) in Catharanthus roseuscambial meristem cells (CMCs) jointly treated with 0.25 mM vindoline and methyl jasmonate (MeJA), suggesting that C. roseus CMCs contain a complete set of the enzymes which are in response to convert vindoline into vincristine and vinblastine. Based on the facts that the transcript levels of vindoline-biosynthetic genes (STR, SGD and D4H) were up-regulated instead of being down-regulated by adding itself to the culture, and that the transcriptional factor ORCA3 was up-regulated simultaneously, we further confirmed that the transcription of STR, SGD, D4H was manipulated by ORCA3.

Topics: Acetates; Antineoplastic Agents, Phytogenic; Cambium; Catharanthus; Cells, Cultured; Cyclopentanes; Gene Expression Regulation, Plant; Oxylipins; Vinblastine; Vincristine

The effects of light and methyl jasmonate (MJ) on the transcription of biosynthetic genes as well as the accumulation of vindoline and catharanthine in Catharanthus roseus C20hi cell suspensions were studied. t16h (the gene encoding tabersonine 16-hydroxylase) could be induced by light and MJ, whereas d4h (the gene encoding deacetoxyvindoline 4-hydroxylase) could only be induced by light. Quantification by UPLC-MS showed that light significantly increased vindoline production in C20hi cells by about 0.49 - 5.51-fold more than that in controls, with the highest yield being 75.3 ng/g of dry weight. The biosynthesis of vindoline was further enhanced by combining MJ with light. The accumulation of catharanthine was not improved by either light or MJ elicitation. These results suggested that light and MJ could promote vindoline, but not catharanthine accumulation in C20hi cells.

Topics: Acetates; Base Sequence; Catharanthus; Cell Line; Chromatography, High Pressure Liquid; Cyclopentanes; DNA Primers; Light; Oxylipins; Polymerase Chain Reaction; Spectrometry, Mass, Electrospray Ionization; Tandem Mass Spectrometry; Vinblastine

Methyl jasmonate, a chemical inducer of secondary metabolism, has been shown to promote vindoline biosynthesis in developing seedlings, as a result of induction of tryptophan decarboxylase (TDC) and desacetylvindoline 4-hydroxylase (D4H). The present studies suggest that jasmonate-based induction of TDC and D4H activities involves modulation of transcriptional, post-transcriptional and post-translational controls. The effects of jasmonate on both enzymes were transient with maximum TDC activity appearing 12 h earlier than that of D4H. Jasmonate treatment of etiolated seedlings neither enhanced TDC activity nor could it replace the light requirement for D4H induction. Jasmonate, therefore, appears to modulate events which are already triggered by developmental and environmental specific controls. Salicylic acid, another chemical inducer of secondary metabolism, was ineffective in activating either TDC or D4H under the experimental conditions used.

Topics: Acetates; Aromatic-L-Amino-Acid Decarboxylases; Cyclopentanes; Enzyme Induction; Light; Mixed Function Oxygenases; Oxylipins; Plant Growth Regulators; Plant Proteins; Plants, Medicinal; Salicylates; Salicylic Acid; Vinblastine