Overexpression of SmbHLH148 induced biosynthesis of tanshinones as well as phenolic acids in Salvia miltiorrhiza hairy roots
SmbHLH148 activated the whole biosynthetic pathways of phenolic acids and tanshinones, thus upregulated the production of both the two groups of pharmaceutical ingredients in Salvia miltiorrhiza.
Phenolic acids and tanshinones are the two important groups of pharmaceutical ingredients presented in Salvia miltiorrhiza Bunge. The bHLH transcription factors could regulate secondary metabolism efficiently in plants. However, there are only some MYCs have been studied on regulation of either phenolic acids or tanshinones biosynthesis. In this study, a bHLH TF named SmbHLH148, which is homologous to AtbHLH148, AtbHLH147 and CubHLH1, was isolated and functionally characterized from S. miltiorrhiza. Transcription of SmbHLH148 could be intensely induced by ABA and also be moderately induced by MeJA and GA. SmbHLH148 is present in all the six tissues and mostly expressed in fibrous root and flowers. Subcellular localization analysis found that SmbHLH148 was localized in the nucleus. Overexpression of SmbHLH148 significantly increased not only three phenolic acids components accumulation but also three tanshinones content. Content of caffeic acid, rosmarinic acid and salvianolic acid B were reached to 2.87-, 4.00- and 5.99-fold of the control in the ObHLH148-3, respectively. Content of dihydrotanshinone I, cryptotanshinone, and tanshinone I were also present highest in ObHLH148-3, reached 2.5-, 5.04- and 3.97-fold of the control, respectively. Expression analysis of pathway genes of phenolic acids and tanshinones in transgenic lines showed that most of them were obviously upregulated. Moreover, transcription of AREB and JAZs were also induced in SmbHLH148 overexpression lines. These results suggested that SmbHLH148 might be taken part in ABA and MeJA signaling and activated almost the whole biosynthetic pathways of phenolic acids and tanshinones, thus the production of phenolic acids and tanshinones were upregulated.
KeywordsbHLH transcription factor Phenolic acids Tanshinones Transgenic Secondary metabolism Salvia miltiorrhiza
This work was supported by the National Natural Science Foundation of China (Nos. 81773835 and 81673536), Natural Science Foundation of Zhejiang Provincial (No. LZ16H280001). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.
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Conflict of interest
The authors declare that they have no conflict of interest.
- Cheng QQ, Su P, Hu YT, He YF, Gao W, Huang LQ (2014) RNA interference-mediated repression of SmCPS (copalyldiphosphate synthase) expression in hairy roots of Salvia miltiorrhiza causes a decrease of tanshinones and sheds light on the functional role of SmCPS. Biotechnol Lett 36:363–369CrossRefGoogle Scholar
- Endo T, Fujii H, Sugiyama A, Nakano M, Nakajima N, Ikoma Y, Omura M, Shimada T (2016) Overexpression of a citrus basic helix-loop-helix transcription factor (CubHLH1), which is homologous to Arabidopsis activation-tagged bri1 suppressor 1 interacting factor genes, modulates carotenoid metabolism in transgenic tomato. Plant Sci 243:35–48CrossRefGoogle Scholar
- Mannen K, Matsumoto T, Takahashi S, Yamaguchi Y, Tsukagoshi M, Sano R, Suzuki H, Sakurai N, Shibata D, Koyama T, Nakayama T (2014) Coordinated transcriptional regulation of isopentenyl diphosphate biosynthetic pathway enzymes in plastids by phytochrome-interacting factor 5. Biochem Bioph Res Co 443:768–774CrossRefGoogle Scholar
- Yamamura C, Mizutani E, Okada K, Nakagawa H, Fukushima S, Tanaka A, Maeda S, Kamakura T, Yamane H, Takatsuji H, Mori M (2015) Diterpenoid phytoalexin factor, a bHLH transcription factor, plays a central role in the biosynthesis of diterpenoid phytoalexins in rice. Plant J 84:1100–1113CrossRefGoogle Scholar
- Zhang H, Hedhili S, Montiel G, Zhang Y, Chatel G, Pre M, Gantet P, Memelink J (2011) The basic helix-loop-helix transcription factor CrMYC2 controls the jasmonate-responsive expression of the ORCA genes that regulate alkaloid biosynthesis in Catharanthus roseus. Plant J 67:61–71CrossRefGoogle Scholar