A candidate gene identified in converting platycoside E to platycodin D from Platycodon grandiflorus by transcriptome and main metabolites analysis

AbstractPlatycodin D and platycoside E are two triterpenoid saponins in Platycodon grandiflorus, differing only by two glycosyl groups structurally. Studies have shown β-Glucosidase from bacteria can convert platycoside E to platycodin D, indicating the potential existence of similar enzymes in P. grandiflorus. An L9(34) orthogonal experiment was performed to establish a protocol for calli induction as follows: the optimal explant is stems with nodes and the optimum medium formula is MS + NAA 1.0 mg/L + 6-BA 0.5 mg/L to obtain callus for experimental use. The platycodin D, platycoside E and total polysaccharides content between callus and plant organs varied wildly. Platycodin D and total polysaccharide content of calli was found higher than that of leaves. While, platycoside E and total polysaccharide content of calli was found lower than that of leaves. Associating platycodin D and platycoside E content with the expression level of genes involved in triterpenoid saponin biosynthesis between calli and leaves, three contigs were screened as putative sequences of β-Glucosidase gene converting platycoside E to platycodin D. Besides, we inferred that some transcription factors can regulate the expression of key enzymes involved in triterpernoid saponins and polysaccharides biosynthesis pathway of P. grandiflorus. Totally, a candidate gene encoding enzyme involved in converting platycoside E to platycodin D, and putative genes involved in polysaccharide synthesis in P. grandiflorus had been identified. This study will help uncover the molecular mechanism of triterpenoid saponins biosynthesis in P. grandiflorus.

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Full-Length Transcriptome Sequences by A Combination of Sequencing Platforms Applied to Isoflavonoid and Triterpenoid Saponin Biosynthesis of Astragalus Mongholicus Bunge

10.21203/rs.3.rs-127368/v1 ◽

2020 ◽

Author(s):

Minzhen Yin ◽

Shanshan Chu ◽

Tingyu Shan ◽

Liangping Zha ◽

Huasheng Peng

Keyword(s):

Single Molecule ◽

Molecular Genetic ◽

Gene Families ◽

Triterpenoid Saponin ◽

Smrt Sequencing ◽

Triterpenoid Saponins ◽

Saponin Biosynthesis ◽

Medicinal Value ◽

Long Time ◽

Sequencing Platforms

Abstract Background: Astragalus mongholicus Bunge is an important medicinal plant and has been used in traditional Chinese medicine for a long history, which is rich in isoflavonoids and triterpenoid saponins. Although these active constituents in A. mongholicus have been discovered for a long time, the molecular genetic basis of the isoflavonoid and triterpenoid saponin biosynthesis pathways is virtually unknown due to the lack of a reference genome. The combination of next-generation sequencing (NGS) and single-molecule real-time (SMRT) sequencing to analyze genes involved in the biosynthetic pathways of secondary metabolites in medicinal plants has been widely recognized.Results: In this study, NGS, SMRT sequencing, and targeted compounds were combined to investigate the association between isoflavonoids and triterpenoid saponins and gene expression in roots, stems and leaves of A. mongholicus. A total of four main isoflavonoids and four astragalosides (belong to triterpenoid saponins) were measured, and 44 differentially expressed genes (DEGs) of nine gene families, 44 DEGs of 16 gene families that encode for enzymes involved in isoflavonoid and triterpenoid saponin biosynthesis were identified, separately. Additionally, transcription factors (TFs) associated with isoflavonoid and triterpenoid saponin biosynthesis were analyzed, including 72 MYBs, 53 bHLHs, 64 AP2-EREBPs and 11 bZIPs. The above transcripts exhibit different expression trends in different organs.Conclusions: Our study provides important genetic information for the essential genes of isoflavonoid and triterpenoid saponin biosynthesis in A. mongholicus, and provides a basis for developing its medicinal value.

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Comparative transcriptome analysis of root, stem, and leaf tissues of Entada phaseoloides reveals potential genes involved in triterpenoid saponin biosynthesis

BMC Genomics ◽

10.1186/s12864-020-07056-1 ◽

2020 ◽

Vol 21 (1) ◽

Author(s):

Weifang Liao ◽

Zhinan Mei ◽

Lihong Miao ◽

Pulin Liu ◽

Ruijie Gao

Keyword(s):

Transcriptome Analysis ◽

Biosynthetic Pathway ◽

Triterpenoid Saponin ◽

Uridine Diphosphate ◽

Comparative Transcriptome ◽

Triterpenoid Saponins ◽

Comparative Transcriptome Analysis ◽

Saponin Biosynthesis ◽

Leaf Tissues ◽

Entada Phaseoloides

Abstract Background Entada phaseoloides (L.) Merr. is an important traditional medicinal plant. The stem of Entada phaseoloides is popularly used as traditional medicine because of its significance in dispelling wind and dampness and remarkable anti-inflammatory activities. Triterpenoid saponins are the major bioactive compounds of Entada phaseoloides. However, genomic or transcriptomic technologies have not been used to study the triterpenoid saponin biosynthetic pathway in this plant. Results We performed comparative transcriptome analysis of the root, stem, and leaf tissues of Entada phaseoloides with three independent biological replicates and obtained a total of 53.26 Gb clean data and 116,910 unigenes, with an average N50 length of 1218 bp. Putative functions could be annotated to 42,191 unigenes (36.1%) based on BLASTx searches against the Non-redundant, Uniprot, KEGG, Pfam, GO, KEGG and COG databases. Most of the unigenes related to triterpenoid saponin backbone biosynthesis were specifically upregulated in the stem. A total of 26 cytochrome P450 and 17 uridine diphosphate glycosyltransferase candidate genes related to triterpenoid saponin biosynthesis were identified. The differential expressions of selected genes were further verified by qPT-PCR. Conclusions The dataset reported here will facilitate the research about the functional genomics of triterpenoid saponin biosynthesis and genetic engineering of Entada phaseoloides.

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Triterpenoid Saponins Isolated fromPlatycodon grandiflorumInhibit Hepatitis C Virus Replication

Evidence-based Complementary and Alternative Medicine ◽

10.1155/2013/560417 ◽

2013 ◽

Vol 2013 ◽

pp. 1-11 ◽

Cited By ~ 10

Author(s):

Jong-Woo Kim ◽

Sang Jin Park ◽

Jong Hwan Lim ◽

Jae Won Yang ◽

Jung Cheul Shin ◽

...

Keyword(s):

Hepatitis C Virus ◽

Hepatitis C ◽

Root Extract ◽

Triterpenoid Saponin ◽

Active Components ◽

Resistant Variant ◽

Triterpenoid Saponins ◽

Platycodin D ◽

Infected Cells ◽

Resistant Strains

Hepatitis C virus (HCV) infection is a major cause of liver disease, including cirrhosis and hepatocellular carcinoma. Due to significant adverse effects and emergence of resistant strains of currently developed anti-HCV agents, plant extracts have been considered to be potential sources of new bioactive compounds against HCV. The aim of this study was to evaluate the functional effects of triterpenoid saponins contained in the root extract ofPlatycodon grandiflorum(PG) on viral enzyme activities and replication in both HCV replicon cells and cell culture grown HCV- (HCVcc-) infected cells. Inhibitory activities of triterpenoid saponins from PG were verified by NS5B RNA-dependent RNA polymerase assay and were further confirmed in the context of HCV replication. Six triterpenoid saponins (platycodin D, platycodin D2, platycodin D3, deapioplatycodin D, deapioplatycodin D2, and platyconic acid A), PG saponin mixture (PGSM), were identified as active components exerting anti-HCV activity. Importantly, PGSM exerted synergistic anti-HCV activity in combination with either interferon-αor NS5A inhibitors. We demonstrated that combinatorial treatment of PGSM and IFN-αefficiently suppressed colony formation with significant reduction in drug resistant variant of HCV. These data suggest that triterpenoid saponin may represent a novel anti-HCV therapeutic agent.

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Comparative transcriptome analysis of root, stem, and leaf tissues of Entada phaseoloides reveals potential genes involved in triterpenoid saponin biosynthesis

10.21203/rs.2.20018/v1 ◽

2020 ◽

Author(s):

Weifang Liao ◽

Lihong Miao ◽

Pulin Liu ◽

Ruijie Gao ◽

Zhinan Mei

Keyword(s):

Transcriptome Analysis ◽

Biosynthetic Pathway ◽

Triterpenoid Saponin ◽

Uridine Diphosphate ◽

Comparative Transcriptome ◽

Triterpenoid Saponins ◽

Comparative Transcriptome Analysis ◽

Saponin Biosynthesis ◽

Leaf Tissues ◽

Entada Phaseoloides

Abstract Background Entada phaseoloides (L.) Merr. is an important traditional medicinal plant. The stem of Entada phaseoloides is popularly used as traditional medicine because of its significance in dispelling wind and dampness and remarkable anti-inflammatory activities. Triterpenoid saponins are the major bioactive compounds of Entada phaseoloides. However, genomic or transcriptomic technologies have not been used to study the triterpenoid saponin biosynthetic pathway in this plant.Results We performed comparative transcriptome analysis of the root, stem, and leaf tissues of Entada phaseoloides with three independent biological replicates and obtained a total of 53.26 Gb clean data and 116,910 unigenes, with an average N50 length of 1218 bp. Putative functions could be annotated to 42,191 unigenes (36.1%) based on BLASTx searches against the Non-redundant, Uniprot, KEGG, Pfam, GO, KEGG and COG databases. Most of the unigenes related to triterpenoid saponin backbone biosynthesis were specifically upregulated in the stem. A total of 26 cytochrome P450 and 17 uridine diphosphate glycosyltransferase candidate genes related to triterpenoid saponin biosynthesis were identified. The differential expressions of selected genes were further verified by qPT-PCR.Conclusions The dataset reported here will facilitate the research about the functional genomics of triterpenoid saponin biosynthesis and genetic engineering of Entada phaseoloides .

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Full-Length Transcriptome Analyses of Genes Involved in Triterpenoid Saponin Biosynthesis of Psammosilene tunicoides Hairy Root Cultures With Exogenous Salicylic Acid

Frontiers in Genetics ◽

10.3389/fgene.2021.657060 ◽

2021 ◽

Vol 12 ◽

Author(s):

Lingye Su ◽

Shufang Li ◽

Hanhan Qiu ◽

Hongfeng Wang ◽

Congcong Wang ◽

...

Keyword(s):

Salicylic Acid ◽

Real Time ◽

Hairy Root ◽

Hairy Roots ◽

Consensus Sequence ◽

Full Length ◽

Triterpenoid Saponin ◽

Rna Seq ◽

Triterpenoid Saponins ◽

Saponin Biosynthesis

Triterpenoid saponins constitute a diverse class of bioactive compounds in medicinal plants. Salicylic acid (SA) is an efficient elicitor for secondary metabolite production, but a transcriptome-wide regulatory network of SA-promoted triterpenoid saponin biosynthesis remains little understood. In the current study, we described the establishment of the hairy root culture system for Psammosilene tunicoides, a triterpenoid saponin-producing medicinal herb in China, using genetic transformation by Agrobacterium rhizogenes. Compared to controls, we found that total saponin content was dramatically increased (up to 2.49-fold) by the addition of 5 mg/L SA in hairy roots for 1 day. A combination of single-molecule real-time (SMRT) and next-generation sequencing (Illumina RNA-seq) was generated to analyze the full-length transcriptome data for P. tunicoides, as well as the transcript profiles in treated (8 and 24 h) and non-treated (0 h) groups with 5 mg/L SA in hairy roots. A total of 430,117 circular consensus sequence (CCS) reads, 16,375 unigenes and 4,678 long non-coding RNAs (lncRNAs) were obtained. The average length of unigenes (2,776 bp) was much higher in full-length transcriptome than that derived from single RNA-seq (1,457 bp). The differentially expressed genes (DEGs) were mainly enriched in the metabolic process. SA up-regulated the unigenes encoding SA-binding proteins and antioxidant enzymes in comparison with controls. Additionally, we identified 89 full-length transcripts encoding enzymes putatively involved in saponin biosynthesis. The candidate transcription factors (WRKY, NAC) and structural genes (AACT, DXS, SE, CYP72A) might be the key regulators in SA-elicited saponin accumulation. Their expression was further validated by quantitative real-time PCR (qRT-PCR). These findings preliminarily elucidate the regulatory mechanisms of SA on triterpenoid saponin biosynthesis in the transcriptomic level, laying a foundation for SA-elicited saponin augmentation in P. tunicoides.

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Key Enzymes of Triterpenoid Saponin Biosynthesis and the Induction of Their Activities and Gene Expressions in Plants

Natural Product Communications ◽

10.1177/1934578x1000500736 ◽

2010 ◽

Vol 5 (7) ◽

pp. 1934578X1000500 ◽

Cited By ~ 4

Author(s):

Chang Ling Zhao ◽

Xiu Ming Cui ◽

Yan Ping Chen ◽

Quan Liang

Keyword(s):

Medicinal Plants ◽

Higher Plants ◽

Squalene Epoxidase ◽

Signal Pathways ◽

Triterpenoid Saponin ◽

Active Components ◽

Gene Expressions ◽

Triterpenoid Saponins ◽

Key Enzymes ◽

Saponin Biosynthesis

Triterpenoid saponins are one of the key active components of many medicinal plants. The biosynthetic pathway of triterpenoid saponins in higher plants and a lot of experimental results both indicated that the key enzymes involved in triterpenoid saponin synthesis are squalene synthase (SS), squalene epoxidase (SE), lupeol synthase (LS), dammarenediol synthase (DS), β-amyrin synthase (β-AS), cytochrome P450-dependent monooxygenase (PDMO), and glycosyltransferase (GT). The activities and coding genes of the key enzymes could be induced by a range of factors in various plant species. However, the effects of the factors on the content and composition of the triterpenoid saponins in specific plants are not certainly coincident, and different factors appear to induce the gene expressions of the key enzymes by different signal pathways and at different levels. This paper could provide a reference for strengthening the triterpenoid saponin-synthesizing capability of specific medicinal plants at enzyme and/or gene expression levels in order to improve the plants’ commercial values.

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Transcriptome Analysis of Clinopodium chinense (Benth.) O. Kuntze and Identification of Genes Involved in Triterpenoid Saponin Biosynthesis

International Journal of Molecular Sciences ◽

10.3390/ijms20112643 ◽

2019 ◽

Vol 20 (11) ◽

pp. 2643 ◽

Cited By ~ 2

Author(s):

Yuanyuan Shi ◽

Shengxiang Zhang ◽

Daiyin Peng ◽

Chenkai Wang ◽

Derui Zhao ◽

...

Keyword(s):

Profile Analysis ◽

Triterpenoid Saponin ◽

Quantitative Real Time Pcr ◽

Pharmacological Activities ◽

Triterpenoid Saponins ◽

The Public ◽

Saponin Biosynthesis ◽

Major Class ◽

Expression Profile Analysis ◽

Transcriptome Database

Clinopodium chinense (Benth.) O. Kuntze (C. chinense) is an important herb in traditional Chinese medicine. Triterpenoid saponins are a major class of active compounds in C. chinense with broad pharmacological activities and hemostatic, antitumor, and anti-hyperglycemic effects. To identify genes involved in triterpenoid saponin biosynthesis, transcriptomic analyses of leaves, stems, and roots from C. chinense were performed. A total of 135,968 unigenes were obtained by assembling the leaf, stem, and root transcripts, of which 102,154 were annotated in public databases. Differentially expressed genes were determined based on expression profile analysis and analyzed for differential expression of unique genes related to triterpenoid saponin biosynthesis. Multiple unigenes encoding crucial enzymes or transcription factors involved in triterpenoid saponin synthesis were identified and analyzed. The expression levels of unigenes encoding enzymes were experimentally validated using quantitative real-time PCR. This study greatly broadens the public transcriptome database for this species and provides a valuable resource for identifying candidate genes involved in the biosynthesis of triterpenoid saponins and other secondary metabolites.

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SMRT- and Illumina‑based RNA‑seq Analyses Unveil Triterpenoid Saponin Biosynthesis and Related Key Genes in Platycodon Grandiflorus (Jacq.) A. DC.

10.21203/rs.3.rs-139609/v1 ◽

2021 ◽

Author(s):

Hanwen Yu ◽

Mengli Liu ◽

Minzhen Yin ◽

Tingyu Shan ◽

Huasheng Peng ◽

...

Keyword(s):

Real Time ◽

Single Molecule ◽

Molecular Mechanisms ◽

Total Content ◽

Triterpenoid Saponin ◽

Triterpenoid Saponins ◽

Sequencing Technologies ◽

Saponin Biosynthesis ◽

Genes Encoding ◽

Key Genes

Abstract Background: Platycodon grandiflorus, a traditional Chinese medicine, contains considerable triterpene saponins with broad pharmacological activities. To date, information on the molecular mechanism of triterpenoid saponin biosynthesis in P. grandiflorus is limited. Here, single-molecule real-time (SMRT) and next-generation sequencing technologies were combined to comprehensively analyse the transcriptome and unveil triterpenoid saponin biosynthesis in P. grandiflorus.Results: We quantified four saponin monomers in P. grandiflorus, and found that the total content of the four saponins was the highest in the roots and the lowest in the stems and leaves. A total of 173,354 non-redundant transcripts generated from the PacBio platform were successfully annotated to seven functional databases, among which 1,765 transcripts were aligned to the "metabolism of terpenoids and polyketides" pathway in the KEGG database. Three full-length transcripts of β-amyrin synthase (β-AS), the key synthase of the β-amyrin, were identified. Furthermore, a total of 132,610 clean reads of BGISEQ sequences were utilised to explore key genes related to the triterpenoid saponin biosynthetic pathway in P. grandiflorus, and 96 differentially expressed genes (DEGs) involved were selected as candidates. Notably, 9 of the 96 DEGs showed the highest expression in the roots, which were considered key genes for synthesising triterpenoid saponins in P. grandiflorus. Furthermore, 3,469 genes encoding transcription factors (TFs) were identified and classified into 57 TF families, including MYB, bHLH, mTERF, and AP2-EREBP. The expression levels of genes were verified by quantitative real-time PCR.Conclusions: Our reliable transcriptome data provide valuable information on the related biosynthesis pathway and may provide new insights into the molecular mechanisms of triterpenoid saponin biosynthesis in P. grandiflorus.

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Comparative transcriptome analysis of root, stem, and leaf tissues of Entada phaseoloides reveals potential genes involved in triterpenoid saponin biosynthesis

10.21203/rs.2.20018/v2 ◽

2020 ◽

Author(s):

Weifang Liao ◽

Zhinan Mei ◽

Lihong Miao ◽

Pulin Liu ◽

Ruijie Gao

Keyword(s):

Transcriptome Analysis ◽

Biosynthetic Pathway ◽

Triterpenoid Saponin ◽

Uridine Diphosphate ◽

Comparative Transcriptome ◽

Triterpenoid Saponins ◽

Comparative Transcriptome Analysis ◽

Saponin Biosynthesis ◽

Leaf Tissues ◽

Entada Phaseoloides

Abstract Background: Entada phaseoloides (L.) Merr. is an important traditional medicinal plant. The stem of Entada phaseoloides is popularly used as traditional medicine because of its significance in dispelling wind and dampness and remarkable anti-inflammatory activities. Triterpenoid saponins are the major bioactive compounds of Entada phaseoloides. However, genomic or transcriptomic technologies have not been used to study the triterpenoid saponin biosynthetic pathway in this plant.Results: We performed comparative transcriptome analysis of the root, stem, and leaf tissues of Entada phaseoloides with three independent biological replicates and obtained a total of 53.26 Gb clean data and 116,910 unigenes, with an average N50 length of 1218 bp. Putative functions could be annotated to 42,191 unigenes (36.1%) based on BLASTx searches against the Non-redundant, Uniprot, KEGG, Pfam, GO, KEGG and COG databases. Most of the unigenes related to triterpenoid saponin backbone biosynthesis were specifically upregulated in the stem. A total of 26 cytochrome P450 and 17 uridine diphosphate glycosyltransferase candidate genes related to triterpenoid saponin biosynthesis were identified. The differential expressions of selected genes were further verified by qPT-PCR. Conclusions: The dataset reported here will facilitate the research about the functional genomics of triterpenoid saponin biosynthesis and genetic engineering of Entada phaseoloides.

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Full-length transcriptome sequences by a combination of sequencing platforms applied to isoflavonoid and triterpenoid saponin biosynthesis of Astragalus mongholicus Bunge

Plant Methods ◽

10.1186/s13007-021-00762-1 ◽

2021 ◽

Vol 17 (1) ◽

Author(s):

Minzhen Yin ◽

Shanshan Chu ◽

Tingyu Shan ◽

Liangping Zha ◽

Huasheng Peng

Keyword(s):

Single Molecule ◽

Gene Families ◽

Specific Gene ◽

Triterpenoid Saponin ◽

Smrt Sequencing ◽

Triterpenoid Saponins ◽

Saponin Biosynthesis ◽

Medicinal Value ◽

Saponin Content ◽

Sequencing Platforms

Abstract Background Astragalus mongholicus Bunge is an important medicinal plant used in traditional Chinese medicine. It is rich in isoflavonoids and triterpenoid saponins. Although these active constituents of A. mongholicus have been discovered for a long time, the genetic basis of isoflavonoid and triterpenoid saponin biosynthesis in this plant is virtually unknown because of the lack of a reference genome. Here, we used a combination of next-generation sequencing (NGS) and single-molecule real-time (SMRT) sequencing to identify genes involved in the biosynthetic pathway of secondary metabolites in A. mongholicus. Results In this study, NGS, SMRT sequencing, and targeted compound analysis were combined to investigate the association between isoflavonoid and triterpenoid saponin content, and specific gene expression in the root, stem, and leaves of A. mongholicus. Overall, 643,812 CCS reads were generated, yielding 121,107 non-redundant transcript isoforms with an N50 value of 2124 bp. Based on these highly accurate transcripts, 104,756 (86.50%) transcripts were successfully annotated by any of the seven databases (NR, NT, Swissprot, KEGG, KOG, Pfam and GO). Levels of four isoflavonoids and four astragalosides (triterpenoid saponins) were determined. Forty-four differentially expressed genes (DEGs) involved in isoflavonoid biosynthesis and 44 DEGs from 16 gene families that encode enzymes involved in triterpenoid saponin biosynthesis were identified. Transcription factors (TFs) associated with isoflavonoid and triterpenoid saponin biosynthesis, including 72 MYBs, 53 bHLHs, 64 AP2-EREBPs, and 11 bZIPs, were also identified. The above transcripts showed different expression trends in different plant organs. Conclusions This study provides important genetic information on the A. mongholicus genes that are essential for isoflavonoid and triterpenoid saponin biosynthesis, and provides a basis for developing the medicinal value of this plant.

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