Yeast Elicitor Research Articles

Phenylpropanoid biosynthetic gene expression and nutrient uptake kinetics for enhanced rosmarinic acid production in suspension cultured cells of Halodule pinifolia

This study investigated the effect of precursors (phenylalanine and tyrosine) and elicitors (methyl jasmonate and yeast) on enhanced rosmarinic acid (RA) biosynthesis, their transcript levels (HpPAL, HpC4H, Hp4CL, HpTAT and HpHPPR) and nutrient uptake kinetics from cell cultures of Halodule pinifolia (Miki) Hartog under in-vitro conditions. Different concentrations of precursors and elicitors were used, among which methyl jasmonate (20 μM) induced higher accumulation of RA (123.16 ± 3.30 mg g−1 DW) followed by phenylalanine (57.086 ± 1.01 mg g−1 DW), tyrosine (28.723 ± 1.55 mg g−1 DW) and yeast elicitor (1.95 mg g−1 DW). Further, the transcript levels of phenylpropanoid biosynthetic genes HpPAL, HpC4H and Hp4CL from MeJA treated cells were found to increase transiently, while TAT and HPPR did not express. It clearly conveyed that H. pinifola culture has undergone phenylpropanoid pathway for RA biosynthesis as compared to the control. Nutrient uptake kinetics determined the consumption of major nutrients such as phosphate, nitrate, sucrose and variations in pH during RA biosynthesis and the optimum pH was found to be pH -7.4, where the concentration of RA accumulated higher during the aforementioned pH. Therefore, the study concludes that methyl jasmonate acts as a potent elicitor for RA accumulation and the enhanced RA biosynthesis in the treated cells might be due to the activation of phenylproponoid genes HpPAL, HpC4H and Hp4CL. Furthermore, the work should be made to influence and optimize the H. pinifolia culture system for better productivity of rosmarinic acid for the food, cosmetic, pharmacy and neutraceutical applications.

CroFGD: Catharanthus roseus Functional Genomics Database.

Catharanthus roseus is a medicinal plant, which can produce monoterpene indole alkaloid (MIA) metabolites with biological activity and is rich in vinblastine and vincristine. With release of the scaffolded genome sequence of C. roseus, it is necessary to annotate gene functions on the whole-genome level. Recently, 53 RNA-seq datasets are available in public with different tissues (flower, root, leaf, seedling, and shoot) and different treatments (MeJA, PnWB infection and yeast elicitor). We used in-house data process pipeline with the combination of PCC and MR algorithms to construct a co-expression network exploring multi-dimensional gene expression (global, tissue preferential, and treat response) through multi-layered approaches. In the meanwhile, we added miRNA-target pairs, predicted PPI pairs into the network and provided several tools such as gene set enrichment analysis, functional module enrichment analysis, and motif analysis for functional prediction of the co-expression genes. Finally, we have constructed an online croFGD database (http://bioinformatics.cau.edu.cn/croFGD/). We hope croFGD can help the communities to study the C. roseus functional genomics and make novel discoveries about key genes involved in some important biological processes.

Effect of Yeast Elicitor on the Enhancement of Kaempferol from in vivo and in vitro Callus Cultures of Dregea volubilis Benth

Effect of Yeast Elicitor on the Enhancement of Kaempferol from in vivo and in vitro Callus Cultures of Dregea volubilis Benth

INFLUENCE OF YEAST EXTRACT ON GENE EXPRESSION OF CRWRKY1 AS A REGULATORY GENE IN ALKALOIDS PATHWAY IN CATHARANTHUS ROSEUS (L.) G. DON

Catharanthus roseus (L.) G. Don is a medicinal plant rich in many alkaloids that are used in treatment of many diseases. It is unique in the production of vinblastine and vincristine compounds, which are used in treatment of several cancers. There are many genes in the biosynthesis pathway that produce these anti-cancer compounds, including the gene Crwrky1 which is a regulatory gene that codes for the transcription factor in the monoterpene indole alkaloid (MIA) pathway. In this study, the effect of yeast extract on crwrky1 gene expression levels from treated calli was studied. Quantitative RT-PCR (QRT-PCR) using SYBR Green I/ROX was used to analyze the changes in the expression level of this gene in response to different treatments of yeast extract. Crwrky1 expression increased to 3.6 folds in treated callus obtained under 0.4 mg/l yeast elicitor for 4 hours (YE2) treatment to compare with the control (untreated) callus.

Chitosan and yeast elicitor in suppressing seed-borne fungi of cucurbitaceous vegetables

Experiments were conducted under laboratory condition to examine the efficacy of Chitosan and Yeast Elicitor to suppress the growth of seed-borne fungi of cucurbitaceous vegetables. Seeds of bottle gourd, sweet gourd, snake gourd, wax gourd and cucumber were collected from seed traders of Mymensingh districts and different seed borne fungi were isolated, purified and identified. Fourteen fungal species belonging to twelve genera consisting of Aspergillus flavus, Aspergillus niger, Botrytis cinerea, Fusarium moniliforme, Fusarium oxysporum, Phoma exigua, Rhizopus stolonifer, Macrophomina phaseolina, Penicillium spp., Curvularia lunata, Chaetomium spp., Colletotrichum spp., Cercospora spp. and Alternaria alternata were isolated and identified. Four concentrations of Chitosan and Yeast Elicitors solutions (200, 500, 1000 & 2000 ppm) including one positive control Vitavax-200 WP (0.35%) were evaluated for controlling seed-borne fungi. Among the seed treating agents Chitosan (2000 ppm) and Yeast Elicitor (2000 ppm) showed better performance in suppressing the seed-borne fungi. Chitosan (2000 ppm) showed superior performance than Yeast Elicitor (2000 ppm). Results from the present study revealed that application of elicitors as seed treatment is a potential alternative of chemical fungicide for selective vegetables.J. Bangladesh Agril. Univ. 16(2): 187-192, August 2018

The enhancement of kaempferol by yeast elicitor in calluscultures of Oxystelma Esculentum (L.F).

The enhancement of kaempferol by yeast elicitor in calluscultures of Oxystelma Esculentum (L.F).

Revelation and cloning of valinomycin synthetase genes in Streptomyces lavendulae ACR-DA1 and their expression analysis under different fermentation and elicitation conditions

Streptomyces species are amongst the most exploited microorganisms due to their ability to produce a plethora of secondary metabolites with bioactive potential, including several well known drugs. They are endowed with immense unexplored potential and substantial efforts are required for their isolation as well as characterization for their bioactive potential. Unexplored niches and extreme environments are host to diverse microbial species. In this study, we report Streptomyces lavendulae ACR-DA1, isolated from extreme cold deserts of the North Western Himalayas, which produces a macrolactone antibiotic, valinomycin. Valinomycin is a K+ ionophoric non-ribosomal cyclodepsipeptide with a broad range of bioactivities including antibacterial, antifungal, antiviral and cytotoxic/anticancer activities. Production of valinomycin by the strain S. lavendulae ACR-DA1 was studied under different fermentation conditions like fermentation medium, temperature and addition of biosynthetic precursors. Synthetic medium at 10°C in the presence of precursors i.e. valine and pyruvate showed enhanced valinomycin production. In order to assess the impact of various elicitors, expression of the two genes viz. vlm1 and vlm2 that encode components of heterodimeric valinomycin synthetase, was analyzed using RT-PCR and correlated with quantity of valinomycin using LC–MS/MS. Annelid, bacterial and yeast elicitors increased valinomycin production whereas addition of fungal and plant elicitors down regulated the biosynthetic genes and reduced valinomycin production. This study is also the first report of valinomycin biosynthesis by Streptomyces lavendulae.

Molecular characterization of an elicitor-responsive 3-hydroxy-3-methylglutaryl coenzyme A reductase gene involved in oleanolic acid production in cell cultures of Achyranthes bidentata

Achyranthes bidentata contains a rich source of important pharmaceutically active triterpene acids including oleanolic acid (OA) as a major one. 3-Hydroxy-3-methylglutaryl coenzyme A reductase (HMGR) is a key enzyme to provide mevalonate for biosynthesis of triterpene acids. In this study, in order to develop a sustainable source of OA, cell suspension cultures were established from shoot cultures of A. bidentata, and a full length cDNA encoding HMGR (designated as AbHMGR) was cloned and characterized. The cDNA contained 2078 nucleotides with a complete open reading frame of 1593 nucleotides, which was predicted to encode a peptide of 530 amino acids. Expression analysis by real-time PCR revealed that AbHMGR mRNA was abundant in A. bidentata roots, stems and leaves. When cultivated in Murashige and Skoog medium supplemented with 1.5 mg/L 1-naphthlcetic acid (NAA) and 1.5 mg/L 6-benzyladenine (6-BA), cells in suspension culture grew rapidly, yielding OA (100.9 mg/L) after 15 days. OA content in cell cultures was increased under the elicitation of methyl jasmonate (MeJA), yeast elicitor (YE) or cadmium chloride (CdCl2). The ultrahigh production of OA was achieved to 371.8 mg/L, a 5.4-fold of the control after 2-day treatment of 0.2 mM MeJA in the cell cultures. Quantitative real-time PCR analysis showed that AbHMGR was expressed at a higher level under the elicitation of MeJA or YE. Our results suggested that OA production may be the result of the up-regulated expression of AbHMGR under the treatment of various elicitors.

Improvement of grape and wine phenolic content by foliar application to grapevine of three different elicitors: Methyl jasmonate, chitosan, and yeast extract

Phenolic compounds play a key role in grape and wine organoleptic properties, being therefore a key parameter in wine quality. Elicitor application constitutes an interesting field of research since it is indirectly involved in the accumulation of phenolic compounds. The aim of this study was to compare the effect of the application of three different elicitors on both grape and wine phenolic content. Methyl jasmonate, chitosan, and a commercial yeast extract were applied to the canopy at veraison and one week later. Results showed that foliar treatments carried out with methyl jasmonate and yeast extract achieved the best results, increasing grape and wine anthocyanin content when compared to the control. Moreover, the application of the yeast elicitor also enhanced grape stilbene content. In contrast, the chitosan treatment did not have a substantial impact on the phenolic compounds. The results of this study indicate that methyl jasmonate and yeast extract applications could be a simple practice to increase grape and wine phenolic content.

Open Stomata 1 Kinase is Essential for Yeast Elicitor-Induced Stomatal Closure in Arabidopsis.

We recently demonstrated that yeast elicitor (YEL)-induced stomatal closure requires a Ca(2+)-dependent kinase, CPK6. A Ca(2+)-independent kinase, Open Stomata 1 (OST1), is involved in stomatal closure induced by various stimuli including ABA. In the present study, we investigated the role of OST1 in YEL-induced stomatal closure in Arabidopsis using a knock-out mutant, ost1-3, and a kinase-deficient mutant, ost1-2. YEL did not induce stomatal closure or activation of guard cell S-type anion channels in the ost1 mutants unlike in wild-type plants. However, YEL did not increase OST1 kinase activity in wild-type guard cells. The YEL-induced stomatal closure and activation of S-type anion channels were also impaired in a gain-of-function mutant of a clade A type 2C protein phosphatase (ABA INSENSITIVE 1), abi1-1C. In the ost1 mutants like in the wild type, YEL induced H2O2 accumulation, activation of non-selective Ca(2+)-permeable cation (ICa) channels and transient elevations in cytosolic free Ca(2+) concentration ([Ca(2+)]cyt) in guard cells. These results suggest that OST1 kinase is essential for stomatal closure and activation of S-type anion channels induced by YEL and that OST1 is not involved in H2O2 accumulation, ICa channel activation or [Ca(2+)]cyt elevations in guard cells induced by YEL.

Cloning and characterization of an elicitor-responsive gene encoding 3-hydroxy-3-methylglutaryl coenzyme A reductase involved in 20-hydroxyecdysone production in cell cultures of Cyanotis arachnoidea

Cyanotis arachnoidea contains a rich source of bioactive phytoecdysteroids (i.e. analogues of insect steroid hormones). 3-Hydroxy-3-methylglutaryl coenzyme A reductase (HMGR) supplies mevalonate for the synthesis of many secondary metabolites including 20-hydroxyecdysone (20E), one of metabolism-enhancing phytoecdysteroids. In this study, in order to develop a sustainable source of 20E, cell suspension cultures were established from shoot cultures of C. arachnoidea, and a full length cDNA encoding HMGR (designated as CaHMGR) was cloned and characterized. The cDNA contained 2037 nucleotides with a complete open reading frame (ORF) of 1800 nucleotides, which was predicted to encode a peptide of 599 amino acids. Expression analysis by real-time PCR revealed that CaHMGR mRNA was abundant in C. arachnoidea stems, roots and leaves. When cultivated in Murashige & Skoog medium supplemented with 0.2 mg L−1 1-naphthlcetic acid (NAA) and 3.0 mg L−1 6-benzyladenine (6-BA), C. arachnoidea cells in suspension culture grew rapidly, yielding 20E (124.14 μg L−1) after 12 days. The content of 20E in cell cultures elicited by 0.2 mM methyl jasmonate (MeJA), 100 mg L−1 yeast elicitor (YE) or 25 μM AgNO3 was increased 8-, 2-, and 6-fold over the control, respectively. Quantitative real-time PCR analysis showed that CaHMGR was expressed at a higher level under the treatment of MeJA or Ag+ elicitor. Our results suggested that 20E accumulation may be the result of the expression up-regulation of CaHMGR involved in the biosynthesis under the treatment of various elicitors.

A β-Secretase (BACE1)-inhibiting C-Methylrotenoid Induced by Yeast Elicitation in Abronia nana Suspension Cultures

Suspension cultures of Abronia nana were established to produce C-methylisoflavones. Treatment of the A. nana cultures with yeast elicitor induced boeravinone E (1), with maximum induction at 24h after elicitor treatment. Of the biotic and abiotic elicitors tested, yeast extract gave the strongest induction of 1. The IC50 value of 1 against β-secretase (β-amyloid cleaving enzyme-1) was 5.57μM. Other proteases such as trypsin, chymotrypsin, and elastase were not inhibited by concentrations up to 1.0mM, indicating that inhibition of β-secretase was specific. 1 was noncompetitive in Dixon plot, and Ki value was 3.79μM.

A role for ABA in the plants response to chitosan

A role for ABA in the plants response to chitosan

Calcium-Dependent Protein Kinase CPK6 Positively Functions in Induction by Yeast Elicitor of Stomatal Closure and Inhibition by Yeast Elicitor of Light-Induced Stomatal Opening in Arabidopsis

Yeast elicitor (YEL) induces stomatal closure that is mediated by a Ca(2+)-dependent signaling pathway. A Ca(2+)-dependent protein kinase, CPK6, positively regulates activation of ion channels in abscisic acid and methyl jasmonate signaling, leading to stomatal closure in Arabidopsis (Arabidopsis thaliana). YEL also inhibits light-induced stomatal opening. However, it remains unknown whether CPK6 is involved in induction by YEL of stomatal closure or in inhibition by YEL of light-induced stomatal opening. In this study, we investigated the roles of CPK6 in induction by YEL of stomatal closure and inhibition by YEL of light-induced stomatal opening in Arabidopsis. Disruption of CPK6 gene impaired induction by YEL of stomatal closure and inhibition by YEL of light-induced stomatal opening. Activation by YEL of nonselective Ca(2+)-permeable cation channels was impaired in cpk6-2 guard cells, and transient elevations elicited by YEL in cytosolic-free Ca(2+) concentration were suppressed in cpk6-2 and cpk6-1 guard cells. YEL activated slow anion channels in wild-type guard cells but not in cpk6-2 or cpk6-1 and inhibited inward-rectifying K(+) channels in wild-type guard cells but not in cpk6-2 or cpk6-1. The cpk6-2 and cpk6-1 mutations inhibited YEL-induced hydrogen peroxide accumulation in guard cells and apoplast of rosette leaves but did not affect YEL-induced hydrogen peroxide production in the apoplast of rosette leaves. These results suggest that CPK6 positively functions in induction by YEL of stomatal closure and inhibition by YEL of light-induced stomatal opening in Arabidopsis and is a convergent point of signaling pathways for stomatal closure in response to abiotic and biotic stress.

Neither Endogenous Abscisic Acid nor Endogenous Jasmonate Is Involved in Salicylic Acid-, Yeast Elicitor-, or Chitosan-Induced Stomatal Closure inArabidopsis thaliana

Salicylic acid (SA), yeast elicitor (YEL), and chitosan (CHT) induced stomatal closure in Arabidopsis wild-type and aba2-2 plants, induced stomatal closure in fluridon-treated wild-type plants, and induced stomatal closure in aos mutants. These results suggest that neither endogenous abscisic acid nor endogenous jasmonic acid is involved in SA-, YEL-, or CHT-induced stomatal closure.

Two guard cell‐preferential MAPKs, MPK9 and MPK12, regulate YEL signalling in Arabidopsis guard cells

We report that two mitogen-activated protein kinases (MAPKs), MPK9 and MPK12, positively regulate abscisic acid (ABA)-induced stomatal closure in Arabidopsis thaliana. Yeast elicitor (YEL) induced stomatal closure accompanied by intracellular reactive oxygen species (ROS) accumulation and cytosolic free calcium concentration ([Ca(2+) ]cyt ) oscillation. In this study, we examined whether these two MAP kinases are involved in YEL-induced stomatal closure using MAPKK inhibitors, PD98059 and U0126, and MAPK mutants, mpk9, mpk12 and mpk9 mpk12. Both PD98059 and U0126 inhibited YEL-induced stomatal closure. YEL induced stomatal closure in the mpk9 and mpk12 mutants but not in the mpk9 mpk12 mutant, suggesting that a MAPK cascade involving MPK9 and MPK12 functions in guard cell YEL signalling. However, YEL induced extracellular ROS production, intracellular ROS accumulation and cytosolic alkalisation in the mpk9, mpk12 and mpk9 mpk12 mutants. YEL induced [Ca(2+) ]cyt oscillations in both wild type and mpk9 mpk12 mutant. These results suggest that MPK9 and MPK12 function redundantly downstream of extracellular ROS production, intracellular ROS accumulation, cytosolic alkalisation and [Ca(2+) ]cyt oscillation in YEL-induced stomatal closure in Arabidopsis guard cells and are shared with ABA signalling.

A novel cytochrome P450 gene from Catharanthus roseus cell line C20hi: cloning and characterization of expression

An expressed sequence tag (EST) obtained from a subtractive-suppression hybridization cDNA library constructed using Catharanthus roseus cell line C20hi and its parental cell line C20D was used to clone a full-length cytochrome P450 cDNA of cyp71d1. The encoded polypeptide contained 507 amino acids with 39–56% identity to other CYP71D subfamily members at the amino acid level. Expression characteristics of cyp71d1 were determined using semi-quantitative RT-PCR. The cyp71d1 transcript was expressed in all three cell lines with the highest level in the cell line C20hi. In the mature C. roseus plant, the cyp71d1 cDNA was highly expressed in petals, roots and stems, but very weakly expressed in young leaves. Its transcription level increased with the development of flowers. 2,4-D could down-regulate the transcription of cyp71d1, as did KT, but only to a minor degree. Neither light nor yeast elicitor could induce the transcription of cyp71d1.

The effects of elicitation on the expression of key enzyme genes and on production of tropane alkaloids in Anisodus acutangulus plant

AbstractTropane alkaloids (TAs) including hyoscyamine, anisodamine, anisodine and scopolamine are widely used as anticholinergic agents, which act on the parasympathetic nervous system, and their clinical demands are increasing rapidly. Anisodus acutangulus is a solanaceous perennial plant that is endemic to China with good potential for TA production and classified as an endangered species in China. Therefore there is a need to improve the TA production in this resource plant by biology methods. In our research, three different important elicitors including methyl jasmonate (MJ), yeast elicitor (YE) and abscisic acid (ABA) were used to study their effects on TA production, and genes expressions in A. acutangulus plant under each treatment were examined. The results revealed that TA production was induced by all the three treatments. However, its quantity varies with inducers used. The effect induced by MJ and YE were significant, while the effect induced by ABA was not obviously. To our knowledge, this is the first report on the relationship between TA production and genes expression profile induced by different elicitors in cultivated A. acutangulus plant, which should provide some useful information on how to improve TA production with the cultivated A. acutangulus plant in the near future.

Yeast Elicitor-Induced Stomatal Closure and Peroxidase-Mediated ROS Production in Arabidopsis

Yeast elicitor (YEL) induces stomatal closure. We investigated reactive oxygen species (ROS) production, nitric oxide (NO) production and [Ca(2+)](cyt) oscillations to clarify YEL signaling in Arabidopsis guard cells. YEL induced ROS accumulation in guard cells. A peroxidase inhibitor [salicylhydroxamic acid (SHAM)] inhibited the stomatal closure and the ROS accumulation, but neither the atrbohD atrbohF mutation nor an NADPH oxidase inhibitor [diphenylene iodonium chloride (DPI)] had any effect. An NO scavenger [2-(4-carboxyphenyl)-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide (cPTIO)] inhibited the YEL-induced stomatal closure and SHAM abolished NO production. YEL-elicited [Ca(2+)](cyt) oscillations were inhibited by SHAM but not by the atrbohD atrbohF mutation. These results indicate that YEL induces stomatal closure accompanied by ROS production mediated by peroxidases and NO production.

Characterization of the Medicago truncatula cell wall proteome in cell suspension culture upon elicitation and suppression of plant defense

In addition to establishing methods for proteome analysis of cell wall proteins (CWPs) for the model plant Medicago truncatula, this work highlights the presence of several protein classes in cell culture. Using a combination of two-dimensional gel electrophoresis (2D-PAGE) and/or liquid chromatography–tandem mass spectrometry (LC–MS/MS), we established the proteome reference map of M. truncatula cell wall proteins. CWPs extracted from purified cell wall fragments resulted in the identification of 46 (2D-PAGE) and 65 (LC–MS/MS) proteins, respectively, with a total of 111 proteins. The identified proteins are involved in various processes, including cell wall modifications, signaling, defense mechanisms, membrane transport, protein synthesis and processing. Further, we conducted comparative proteome analysis to identify changes in protein composition during interaction of M. truncatula cell suspension culture with a pathogen-derived yeast elicitor (YE) and suppressor using Sinorhizobium meliloti LPS. 2D-PAGE analysis for the CWPs after YE and LPS treatment resembled the proteome map of YE alone, with a few up-regulated proteins involved in defense, and in the case of the LPS-treated cell wall proteome, there was no significant difference observed. Using this approach, proteins involved in defense, such as l-ascorbate peroxidase, specifically targeted proteins to the cell wall during defense, including glyceraldehyde-3-phosphate dehydrogenase (GAPDH), and proteins that play an important role during growth and development were identified. Also, some defense-related proteins were absent in the same gel after YE treatment, suggesting that oxidant protection is regulated by these proteins.