Nitrogen treatment enhances sterols and withaferin A through transcriptional activation of jasmonate pathway, WRKY transcription factors, and biosynthesis genes in Withania somnifera (L.) Dunal

Singh

et al. 2018

Sci Rep

Tissue specific biosynthesis of secondary metabolites is a distinguished feature of medicinal plants. Withania somnifera, source of pharmaceutically important withanolides biosynthesizes withaferin-A in leaves and withanolide-A in roots. To increase the in planta withanolides production, a sustainable approach needs to be explored. Here, we isolated endophytes from different parts of W. somnifera plants and their promising role in in planta withanolide biosynthesis was established in both in-vivo grown as well in in-vitro raised composite W. somnifera plants. Overall, the fungal endophytes improved photosynthesis, plant growth and biomass, and the root-associated bacterial endophytes enhanced the withanolide content in both in-vivo and in-vitro grown plants by modulating the expression of withanolide biosynthesis genes in leaves and roots. Surprisingly, a few indole-3-acetic acid (IAA)-producing and nitrogen-fixing root-associated endophytes could induce the biosynthesis of withaferin-A in roots by inducing in planta IAA-production and upregulating the expression of withanolide biosynthesis genes especially MEP-pathway genes (DXS and DXR) in roots as well. Results indicate the role of endophytes in modulating the synthesis and site of withanolides production and the selected endophytes can be used for enhancing the in planta withanolide production and enriching roots with pharmaceutically important withaferin-A which is generally absent in roots.

Section: Discussionsupporting

confidence: 90%

Endophytes of Withania somnifera modulate in planta content and the site of withanolide biosynthesis

Singh

et al. 2018

Sci Rep

“…In our previous study, nitrogen treatment enhanced the accumulation of phytosterols and withanolides, which were correlated with induced expression of WsWRKY1 (expressed sequence tag (EST) accession number ), indicating the possible role of WsWRKY1 in the regulation of triterpenoids (Pal et al ., ). Here, to further investigate the involvement of WsWRKY1 in the regulation of triterpenoid withanolide biosynthesis, its expression was evaluated in response to MeJA and salicin that are known to induce withanolides accumulation in W. somnifera (Rana et al ., ).…”

Section: Resultsmentioning

confidence: 97%

“…In this study, we set out to investigate the transcriptional regulation of triterpenoid withanolide biosynthesis in W. somnifera . Our previous study found that a particular TF belonging to the WRKY class ( WsWRKY1 ) was highly induced in response to nitrogen treatment, with a corresponding increase in withanolides (withaferin A) as well as phytosterols (Pal et al ., ). Here, a series of investigations was carried out to further determine the functional role of WsWRKY1 .…”

Section: Discussionmentioning

confidence: 97%

“…In our recent study, nitrogen treatment of W. somnifera enhanced the accumulation of phytosterols and withaferin A which correlated with higher expression of sterol pathway genes. Further, it was shown that, among different classes of TFs analyzed (WRKY, MYB, bHLH and bZIP), a particular TF WsWRKY1 ( W. somnifera WRKY1) was highly induced in nitrogen‐treated samples (Pal et al ., ). Here, we present the functional characterization of WsWRKY1 which shows its important role in regulation of triterpenoid biosynthesis and defense in W. somnifera .…”

Section: Introductionmentioning

confidence: 97%

See 1 more Smart Citation

A WRKY transcription factor from Withania somnifera regulates triterpenoid withanolide accumulation and biotic stress tolerance through modulation of phytosterol and defense pathways

et al. 2017

Self Cite

Withania somnifera produces pharmacologically important triterpenoid withanolides that are derived via phytosterol pathway; however, their biosynthesis and regulation remain to be elucidated. A jasmonate- and salicin-inducible WRKY transcription factor from W. somnifera (WsWRKY1) exhibiting correlation with withaferin A accumulation was functionally characterized employing virus-induced gene silencing and overexpression studies combined with transcript and metabolite analyses, and chromatin immunoprecipitation assay. WsWRKY1 silencing resulted in stunted plant growth, reduced transcripts of phytosterol pathway genes with corresponding reduction in phytosterols and withanolides in W. somnifera. Its overexpression elevated the biosynthesis of triterpenoids in W. somnifera (phytosterols and withanolides), as well as tobacco and tomato (phytosterols). Moreover, WsWRKY1 binds to W-box sequences in promoters of W. somnifera genes encoding squalene synthase and squalene epoxidase, indicating its direct regulation of triterpenoid pathway. Furthermore, while WsWRKY1 silencing in W. somnifera compromised the tolerance to bacterial growth, fungal infection, and insect feeding, its overexpression in tobacco led to improved biotic stress tolerance. Together these findings demonstrate that WsWRKY1 has a positive regulatory role on phytosterol and withanolides biosynthesis, and defense against biotic stress, highlighting its importance as a metabolic engineering tool for simultaneous improvement of triterpenoid biosynthesis and plant defense.

“…The expression profiles of these CYPs showed chemotype-specific and tissue-specific variation, as well as variation in response to physiological and developmental factors. To expand the understanding of expression of genes in relation to withanolide biosynthetic pathway, Pal et al ( 2016 ) perform experiments with different concentrations of fertilizers on fresh twigs of W. somnifera . Treated twigs related to highest accumulation of withaferin-A has been selected to analyse expression pattern of CYPs, allene oxide cyclases (AOCs) and few other pathway related genes.…”

Section: Proposed Pathways For Biosynthesis Of Withanolides: Medicinamentioning

confidence: 99%

Withania somnifera: Advances and Implementation of Molecular and Tissue Culture Techniques to Enhance Its Application

et al. 2017

Withania somnifera, commonly known as Ashwagandha an important medicinal plant largely used in Ayurvedic and indigenous medicine for over 3,000 years. Being a medicinal plant, dried powder, crude extract as well as purified metabolies of the plant has shown promising therapeutic properties. Withanolides are the principal metabolites, responsible for the medicinal properties of the plant. Availability and amount of particular withanolides differ with tissue type and chemotype and its importance leads to identification characterization of several genes/ enzymes related to withanolide biosynthetic pathway. The modulation in withanolides can be achieved by controlling the environmental conditions like, different tissue culture techniques, altered media compositions, use of elicitors, etc. Among all the in vitro techniques, hairy root culture proved its importance at industrial scale, which also gets benefits due to more accumulation (amount and number) of withanolides in roots tissues of W. somnifera. Use of media compostion and elicitors further enhances the amount of withanolides in hairy roots. Another important modern day technique used for accumulation of desired secondary metabolites is modulating the gene expression by altering environmental conditions (use of different media composition, elicitors, etc.) or through genetic enginnering. Knowing the significance of the gene and the key enzymatic step of the pathway, modulation in withanolide contents can be achieved upto required amount in therapeutic industry. To accomplish maximum productivity through genetic enginnering different means of Withania transformation methods have been developed to obtain maximum transformation efficiency. These standardized transformation procedues have been used to overexpress/silence desired gene in W. somnifera to understand the outcome and succeed with enhanced metabolic production for the ultimate benefit of human race.