SmMYC complexes cooperative mediate methyl jasmonate regulates tanshinone biosynthesis in Salvia miltiorrhiza

The traditional Chinese medicine Salvia miltiorrhiza is widely used in the treatment of cardiovascular diseases due to its rich of phenolic acids and tanshinones. Among S. miltiorrhiza from different producing areas in China, the Sichuan ecotype (S. m.-SC) has the highest accumulation of phenolic acids, while the lack of tanshinones content seriously restricts its quality and downstream industrial development. It is urgent to develop the quality improvement cultivation technology and explore the regulation mechanism of tanshinone biosynthesis. Herein, we systematically evaluated the interaction between exogenous hormones and tanshinone accumulation in S. m.-SC, and found that Methyl jasmonate (MeJA) could significantly increase the accumulation of tanshinones in S. m.-SC. Based on the integration of pan-transcriptome datasets led to the construction of a tanshinones core metabolic regulatory network and SmMYC3 is a novel discovered positive regulator of tanshinone biosynthesis following SmMYC2a/2b. Yeast one-hybrid (Y1H) and Dual-LUC experiments showed that SmMYC3 directly binds to the G-box elements in the promoters of tanshinone biosynthesis genes (SmGGPPS1, SmKSL1, and SmCYP76AH1). Unexpectedly, SmMYC3-SmMYC2b/SmMYC3 complex identified via yeast two-hybrid (Y2H), Bimolecular fluorescence complementation (Bi-FC) and Dual-LUC experiments has robust enhance the transcriptional activation of the tanshinone metabolism genes upon the involvement of MeJA signaling. This work further expanded the molecular network of MYC transcription factor regulating tanshinone metabolism, clarified the regulatory mechanism of tanshinone biosynthesis mediated by MeJA, and drew a new blueprint for the application of MeJA in the increase of tanshinone content in S. miltiorrhiza.