Abstract
Key message
Overexpression and antisense expression of Sm4CL2 re-directed the biosynthesis of salvianolic acids and tanshinones in Salvia miltiorrhiza hairy roots.
Abstract
Danshen (Salvia miltiorrhiza Bunge) is a widely used traditional Chinese medicine and its main active ingredients are water-soluble phenolic acids and lipophilic diterpenoids which are produced through the phenylpropanoid pathway and terpenoid pathway, respectively. 4-Coumaric acid: Coenzyme A ligase (4CL) is a key enzyme in the phenylpropanoid metabolism. We had obtained Sm4CL2-overexpressing (Sm4CL2-OE) and antisense Sm4CL2-expressing (anti-Sm4CL2) danshen hairy roots over ten years ago. In the follow-up study, we found that total salvianolic acids in Sm4CL2-OE-4 hairy roots increased to 1.35 times of the control-3, and that in anti-Sm4CL2-1 hairy roots decreased to 37.32% of the control-3, but tanshinones in anti-Sm4CL2-1 was accumulated to 1.77 ± 0.16 mg/g of dry weight, compared to undetectable in Sm4CL2-OE-4 and the control-3 hairy roots. Interestingly, Sm4CL2-OE-4 hairy roots contained more lignin, 1.36 times of the control-3, and enhanced cell wall and xylem lignification. Transcriptomic analysis revealed that overexpression of Sm4CL2 caused the upregulation of other phenylpropanoid pathway genes and antisense Sm4CL2 expression resulted in the downregulation of other phenylpropanoid pathway genes but activated the expression of terpenoid pathway genes like SmCYP76AK5, SmGPPS.SSUII.1 and SmDXS2. Protein–protein interaction analysis suggested that Sm4CL2 might interact with PAL, PAL4, CSE, CCoAOMT and SmCYP84A60, and appeared to play a key role in the interaction network. The tracking work in this study proved that Sm4CL2 could redirect both salvianolic acids and tanshinones biosynthesis possibly through synergistically regulating other pathway genes. It also indicated that genetic modification of plant secondary metabolism with biosynthetic gene might cause other responses through protein–protein interactions.
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Acknowledgements
We greatly appreciate Prof. Zhibi Hu and Mrs Jiyan Zhou, Shanghai University of traditional Chinese medicine, for their advice and help.
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This research was funded by the Budgetary items of Shanghai Municipal Education Commission (No. 2021LK113).
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R.T. did most of the experiments, analyzed the experimental data and wrote the draft. M.C. helped to cultivate danshen hairy roots. L.W. guided to analysis results of hairy roots micro section. J.Z. did the quality and quantitative analysis of hairy roots compounds. S. Z. helped to design and guide the experiment and revised the manuscript. All authors read and approved the final manuscript.
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Tan, R., Chen, M., Wang, L. et al. A tracking work on how Sm4CL2 re-directed the biosynthesis of salvianolic acids and tanshinones in Salvia miltiorrhiza hairy roots. Plant Cell Rep 42, 297–308 (2023). https://doi.org/10.1007/s00299-022-02957-9
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DOI: https://doi.org/10.1007/s00299-022-02957-9