The transcription factor PnMYB38 orchestrates methyl jasmonate-induced saponin biosynthesis in Panax notoginseng

Abstract

Panax notoginseng is an important medicinal plant, and saponins are the primary active components that are the key determinants of pharmaceutical quality. Research has indicated that methyl jasmonate (MeJA) enhances the saponins accumulation in P. notoginseng, but the specific MeJA-responsive transcription factors (TFs) that regulate this process remains unidentified. Given the critical role of MYB TFs in the regulation of plant secondary metabolism, this study aimed to elucidate the regulatory mechanisms of the MYB TF family in P. notoginseng under MeJA treatment. Genome-wide screening led to the identification 110 MYB genes, followed by a comprehensive analysis of their phylogenetic relationships, conserved motifs, gene structures, cis-acting elements, chromosomal localization, and collinearity. Integrated transcriptomic and metabolomic analyses showed that MeJA treatment significantly altered the expression patterns of 84 MYB genes while also promoting saponin accumulation in P. notoginseng leaves. Co-expression network analysis revealed a significant correlation between PnMYB38 and saponin metabolites, highlighting the pivotal regulatory function of this TF. Subcellular localization experiments confirmed nuclear localization of PnMYB38. Yeast one-hybrid, electrophoretic mobility shift assay, and dual-luciferase assays demonstrated that PnMYB38 directly and specifically bound to the promoters of key saponin biosynthesis genes ( PnSE and PnDS), thereby inducing their expression. This study comprehensively characterized the functional role of PnMYB38 in regulating MeJA-mediated saponin biosynthesis in P. notoginseng, proposed a ‘MeJA- PnMYB38-saponin biosynthesis’ regulatory network that provided novel insights into the transcriptional regulatory mechanism of saponin biosynthesis, and established a foundation for molecular breeding targeting MYB TFs and metabolic engineering.

Acknowledgements

We thank the editor and anonymous reviewers for providing valuable comments on the manuscript. Y.L. acknowledges financial support from the Major Science and Technology Special Project of Yunnan Province (Grant No. 202202AG050021) and National Science Foundation of China (Grant No. 32360151). The Ability Establishment of Sustainable Use for Valuable Chinese Medicine Resources (Grant No. 2060302). Kunming University of Science and Technology Research Startup Fund (Grant No. KKZ3202560055).

Author contributions

Z.Y. and L.Y. contributed equally to this work. L.Y. and Z.Y. performed data analysis, experiment section, and manuscript writing. Z.Z., M.L., J.H., S.W., H.W., and Q.Y. provided help for experiment section. Y.T. and Y.Y. provided sample materials. X.C., Y.Y., and Y.L. conceived and designed the project, and then revised the manuscript.

Data availability

The RNA-seq data have been deposited at the China National GeneBank (CNGBdb, https://db.cngb.org, Project number: PRJCA048040) and are publicly available as of the date of publication.

Conflicts of interest statement

The authors declare that they have no competing interests.

Supplementary material

Supplementary material is available at Horticulture Research online.

References

[1] Li W, Shi H, Wu X . A narrative review of Panax notoginseng: unique saponins and their pharmacological activities . J Ginseng Res. 2025; 49: 118-33

[2] Wei G, Zhang G, Li M. et al. Panax notoginseng: panoramagram of phytochemical and pharmacological properties, biosynthesis, and regulation and production of ginsenosides . Hortic Res. 2024; 11: uhae170

[3] Hou M, Wang R, Zhao S. et al. Ginsenosides in Panax genus and their biosynthesis. Acta Pharm Sin B. 2021; 11: 1813-34

[4] Kim YK, Kim YB, Uddin MR. et al. Enhanced triterpene accumulation in Panax ginseng hairy roots overexpressing mevalonate-5-pyrophosphate decarboxylase and farnesyl pyrophosphate synthase . ACS Synth Biol. 2014; 3: 773-9

[5] Luo H, Sun C, Sun Y. et al. Analysis of the transcriptome of Panax notoginseng root uncovers putative triterpene saponin-biosynthetic genes and genetic markers . BMC Genomics. 2011; 12: S5

[6] Dong H, Qi X . Biosynthesis of triterpenoids in plants: pathways, regulation, and biological functions. Curr Opin Plant Biol. 2025; 85: 102701

[7] Thimmappa R, Geisler K, Louveau T. et al. Triterpene biosynthesis in plants. Annu Rev Plant Biol. 2014; 65: 225-57

[8] Lu C, Zhao SJ, Feng PC. et al. Functional analysis of the promoter of a UDP-glycosyltransferase gene from Panax quinquefolius . Plant Cell Tissue Organ Cult. 2018; 135: 381-93

[9] Zheng H, Fu X, Shao J. et al. Transcriptional regulatory network of high-value active ingredients in medicinal plants. Trends Plant Sci. 2023; 28: 429-46

[10] Li Y, Li Z, Xu C. et al. WRKYs as regulatory hubs of secondary metabolic networks: diverse inducers and distinct responses. Plant Commun. 2025; 6: 101438

[11] Huang Y, Shi Y, Hu X. et al. PnNAC2 promotes the biosynthesis of Panax notoginseng saponins and induces early flowering . Plant Cell Rep. 2024; 43: 73

[12] Zhao X, Ding J, Ma J. et al. Functional characterization of EsMYB transcription factors in triterpene glycosides biosynthesis pathway in Eleutherococcus senticosus . Ind Crop Prod. 2025; 236: 122046

[13] Lu N, Roldan M, Dixon RA . Characterization of two TT2-type MYB transcription factors regulating proanthocyanidin biosynthesis in tetraploid cotton, Gossypium hirsutum . Planta. 2017; 246: 323-35

[14] Zhang X, Ge F, Deng B. et al. Molecular cloning and characterization of PnbHLH1 transcription factor in Panax notoginseng . Molecules (Basel, Switzerland). 2017; 22: 1268

[15] Zhang D, Zhou H, Zhang Y. et al. Diverse roles of MYB transcription factors in plants. J Integr Plant Biol. 2025; 67: 539-62

[16] Bulanov AN, Andreeva EA, Tsvetkova NV. et al. Regulation of flavonoid biosynthesis by the MYB-bHLH-WDR (MBW) complex in plants and its specific features in cereals. Int J Mol Sci. 2025; 26: 734

[17] Man J, Shi Y, Huang Y. et al. PnMYB4 negatively modulates saponin biosynthesis in Panax notoginseng through interplay with PnMYB1 . Hortic Res. 2023; 10: uhad134

[18] Wu X, Xia M, Su P. et al. MYB transcription factors in plants: a comprehensive review of their discovery, structure, classification, functional diversity and regulatory mechanism. Int J Biol Macromol. 2024; 282: 136652

[19] Xia P, Hu W, Zheng Y. et al. Structural and interactions analysis of a transcription factor PnMYB2 in Panax notoginseng . J Plant Physiol. 2022; 275: 153756

[20] Yang Z, Li Y, Gao F. et al. MYB21 interacts with MYC2 to control the expression of terpene synthase genes in flowers of Freesia hybrida and Arabidopsis thaliana . J Exp Bot. 2020; 71: 4140-58

[21] Du H, Zhai Z, Pu J. et al. Two tandem R2R3 MYB transcription factor genes cooperatively regulate anthocyanin accumulation in potato tuber flesh. Plant Biotechnol J. 2025; 23: 1521-34

[22] Zhou M, Memelink J . Jasmonate-responsive transcription factors regulating plant secondary metabolism. Biotechnol Adv. 2016; 34: 441-9

[23] Jeyasri R, Muthuramalingam P, Karthick K. et al. Methyl jasmonate and salicylic acid as powerful elicitors for enhancing the production of secondary metabolites in medicinal plants: an updated review. Plant Cell Tissue Organ Cult. 2023; 153: 447-58

[24] Deng B, Huang Z, Ge F. et al. An AP2/ERF family transcription factor PnERF1 raised the biosynthesis of saponins in Panax notoginseng . J Plant Growth Regul. 2017; 36: 691-701

[25] Li Y, Lin Y, Jia B. et al. Transcriptome analysis reveals molecular mechanisms underlying methyl jasmonate-mediated biosynthesis of protopanaxadiol-type saponins in Panax notoginseng leaves . J Plant Biol. 2022; 65: 29-41

[26] Liu T, Luo T, Guo X. et al. PgMYB2, a MeJA-responsive transcription factor, positively regulates the dammarenediol synthase gene expression in Panax ginseng . Int J Mol Sci. 2019; 20: 2219

[27] Yuan S, Jiang H, Wang Y. et al. A 3R-MYB transcription factor is involved in methyl jasmonate-induced disease resistance in Agaricus bisporus and has implications for disease resistance in Arabidopsis . J Adv Res. 2025; 73: 117-31

[28] Zhang W, Zhang J, Fan Y. et al. RNA sequencing analysis reveals PgbHLH28 as the key regulator in response to methyl jasmonate-induced saponin accumulation in Platycodon grandiflorus . Hortic Res. 2024; 11: uhae058

[29] Xing B, Qiu L, Wang Z. et al. A novel synergistic regulatory mechanism involving the MYB39-MYB111-bHLH51-TTG1 module in the phenolic and diterpenoid biosynthetic pathways of Salvia miltiorrhiza . Plant Biotechnol J. 2025; 23: 4367-80

[30] Byun JY, Nguyen TT, Cho BK. et al. Rap1 overexpression boosts triterpenoid saponin production in yeast by enhancing precursor supply and heterologous gene expression. Microb Cell Factories. 2025; 24: 47

[31] Kranz HD, Denekamp M, Greco R. et al. Towards functional characterisation of the members of the R2R3- MYB gene family from Arabidopsis thaliana . Plant J. 1998; 16: 263-76

[32] Stracke R, Werber M, Weisshaar B . The R2R3- MYB gene family in Arabidopsis thaliana . Curr Opin Plant Biol. 2001; 4: 447-56

[33] Shen JY, Su TH, Yu D. et al. Evolution by gene duplication: in the era of genomics. Yi chuan. 2025; 47: 147-71

[34] Cannon SB, Mitra A, Baumgarten A. et al. The roles of segmental and tandem gene duplication in the evolution of large gene families in Arabidopsis thaliana . BMC Plant Biol. 2004; 4: 10

[35] Mrudulakumari Vasudevan U, Lee EY . Flavonoids, terpenoids, and polyketide antibiotics: role of glycosylation and biocatalytic tactics in engineering glycosylation. Biotechnol Adv. 2020; 41: 107550

[36] Katiyar A, Smita S, Lenka SK. et al. Genome-wide classification and expression analysis of MYB transcription factor families in rice and Arabidopsis. BMC Genomics. 2012; 13: 544

[37] Liu M, Li K, Sheng S. et al. Transcriptome analysis of MYB transcription factors family and PgMYB genes involved in salt stress resistance in Panax ginseng . BMC Plant Biol. 2022; 22: 479

[38] Song G, Yan Y, Guo C. et al. Identification and expression analysis of R2R3-MYB transcription factors associated with flavonoid biosynthesis in Panax quinquefolius . Int J Mol Sci. 2024; 25: 3709

[39] Yu M, Ma C, Tai B. et al. Unveiling the regulatory mechanisms of nodules development and quality formation in Panax notoginseng using multi-omics and MALDI-MSI . J Adv Res. 2025; 69: 463-75

[40] Xu R, Wang Y, Zheng H. et al. Salt-induced transcription factor MYB74 is regulated by the RNA-directed DNA methylation pathway in Arabidopsis. J Exp Bot. 2015; 66: 5997-6008

[41] Li H, Yao Y, Li X. et al. Comparative genomics analysis of the MYB gene family in barley: preliminary insights into evolution and biological function in Blue Qingke. PeerJ. 2024; 12: e18443

[42] Cao Y, Mei Y, Zhang R. et al. Transcriptional regulation of flavonol biosynthesis in plants. Hortic Res. 2024; 11: uhae043

[43] Chen C, Zhou X, Cao B. et al. The transcription factor MYB8 positively regulates flavonoid biosynthesis of Scutellaria baicalensis in response to drought stress . Plant Cell Environ. 2025; 48: 8898-914

[44] Wei J, Mu X, Wang S. et al. Integrated metabolome and transcriptome analysis provides insights into the mechanisms of terpenoid biosynthesis in tea plants (Camellia sinensis) . Food Res Int. 2025; 201: 115542

[45] Yang J, Xue Q, Li C. et al. A chromosome-level Dendrobium moniliforme genome assembly reveals the regulatory mechanisms of flavonoid and carotenoid biosynthesis pathways . Acta Pharm Sin B. 2025; 15: 2253-72

[46] Zhang D-X, Liu B-Y, Xue F-F. et al. Paclobutrazol induces triterpenoid biosynthesis via downregulation of the negative transcriptional regulator SlMYB in Sanghuangporus lonicericola . Commun Biol. 2025; 8: 551

[47] Chen X, Hudson GA, Mineo C. et al. Deciphering triterpenoid saponin biosynthesis by leveraging transcriptome response to methyl jasmonate elicitation in Saponaria vaccaria . Nat Commun. 2023; 14: 7101

[48] Farag MA, Mekky H, El-Masry S . Metabolomics driven analysis of Erythrina lysistemon cell suspension culture in response to methyl jasmonate elicitation . J Adv Res. 2016; 7: 681-9

[49] Tugizimana F, Ncube EN, Steenkamp PA. et al. Metabolomics-derived insights into the manipulation of terpenoid synthesis in Centella asiatica cells by methyl jasmonate . Plant Biotechnol Rep. 2015; 9: 125-36

[50] Wang J, Gao W, Zuo B. et al. Effect of methyl jasmonate on the ginsenoside content of Panax ginseng adventitious root cultures and on the genes involved in triterpene biosynthesis . Res Chem Intermed. 2013; 39: 1973-80

[51] Li Y, Chen X, Wang J. et al. Two responses to MeJA induction of R2R3-MYB transcription factors regulate flavonoid accumulation in Glycyrrhiza uralensis Fisch . PLoS One. 2020; 15: e0236565

[52] Qiu B, Chen H, Zheng L. et al. An MYB transcription factor modulates Panax notoginseng resistance against the root rot pathogen Fusarium solani by regulating the jasmonate acid signaling pathway and photosynthesis . Phytopathology. 2022; 112: 1323-34

[53] Liu H, Li L, Fu X. et al. A aMYB108 is the core factor integrating light and jasmonic acid signaling to regulate artemisinin biosynthesis in Artemisia annua . New Phytol. 2023; 237: 2224-37

[54] Lin T, Du J, Zheng X. et al. Comparative transcriptome analysis of MeJA-responsive AP2/ERF transcription factors involved in notoginsenosides biosynthesis. 3 Biotech. 2020; 10: 290

[55] Zhao H, Du R, Yl H. et al. Effects of methyl jasmonate and salicylhydroxamic acid on the biosynthesis of flavonoids in Glycyrrhiza glabra L. hairy roots . Plant Stress. 2025; 15: 100774

[56] Li X, Zheng Y, Liu M. et al. Weighted gene co-expression network analysis and identification of ginsenoside biosynthesis candidate genes for ginseng adventitious roots under MeJA treatment. Genes Genomics. 2024; 46: 1473-85

[57] Jia N, Liu J, Sun Y. et al. Citrus sinensis MYB transcription factors CsMYB330 and CsMYB308 regulate fruit juice sac lignification through fine-tuning expression of the Cs4CL1 gene. Plant Sci. 2018; 277: 334-43

[58] Ma D, Reichelt M, Yoshida K. et al. Two R2R3-MYB proteins are broad repressors of flavonoid and phenylpropanoid metabolism in poplar. Plant J. 2018; 96: 949-65

[59] Reddy VA, Wang Q, Dhar N. et al. Spearmint R2R3-MYB transcription factor MsMYB negatively regulates monoterpene production and suppresses the expression of geranyl diphosphate synthase large subunit (MsGPPS.LSU) . Plant Biotechnol J. 2017; 15: 1105-19

[60] Zhang P, Liu X, Yu X. et al. The MYB transcription factor CiMYB42 regulates limonoids biosynthesis in citrus. BMC Plant Biol. 2020; 20: 254

[61] Zheng L, Qiu B, Su L. et al. Panax notoginseng WRKY transcription factor 9 is a positive regulator in responding to root rot pathogen Fusarium solani . Front Plant Sci. 2022; 13: 930644

[62] Li K, Li M, Liu X. et al. Transcriptional regulation modulates saponin biosynthesis of Panax notoginseng response to root knot nematodes . J Appl Res Med Aromat Plants. 2024; 42: 100560

[63] Wen C, Zhang Z, Shi Q. et al. Methyl jasmonate- and salicylic acid-induced transcription factor ZjWRKY18 regulates triterpenoid accumulation and salt stress tolerance in Jujube . Int J Mol Sci. 2023; 24: 3899

[64] Rouster J, Leah R, Mundy J. et al. Identification of a methyl jasmonate-responsive region in the promoter of a lipoxygenase 1 gene expressed in barley grain. Plant J. 1997; 11: 513-23

[65] Cao R, Lv B, Shao S. et al. The SmMYC2-SmMYB36 complex is involved in methyl jasmonate-mediated tanshinones biosynthesis in Salvia miltiorrhiza . Plant J. 2024; 119: 746-61

[66] Li S, Dong Y, Li D. et al. Eggplant transcription factor SmMYB5 integrates jasmonate and light signaling during anthocyanin biosynthesis. Plant Physiol. 2024; 194: 1139-65

[67] Li XJ, Zhou XH, Bao AK . Genome-wide analysis of the R2R3-MYB gene family and identification of candidate genes that regulate isoflavone biosynthesis in red clover (Trifolium pratense) . Int J Biol Macromol. 2024; 282: 137182

[68] Su D, Zheng J, Yi Y. et al. Genome-wide identification of the MYB gene family and screening of potential genes involved in fatty acid biosynthesis in walnut. Phyton Int J Exp Bot. 2024; 93: 2317-37

[69] Yang K, Dong Q, Wu J. et al. Genome-wide analysis of the R2R3-MYB transcription factor gene family expressed in Juglans regia under abiotic and biotic stresses . Ind Crop Prod. 2023; 198: 116709

[70] Wei Q, Chen R, Wei X. et al. Genome-wide identification of R2R3-MYB family in wheat and functional characteristics of the abiotic stress responsive gene TaMYB344. BMC Genomics. 2020; 21: 792

[71] Yang L, Guo J, Wang P. et al. Integrative transcriptomic and metabolomic analysis reveals NBS resistance genes and metabolic pathways of Panax japonicus var. major . Ind Crop Prod. 2024; 216: 118722