Spatiotemporal and Transcriptional Characterization on Tanshinone Initial Synthesis in Salvia miltiorrhiza Roots

Tanshinones are the bioactive constituents of Danshen (Salvia miltiorrhiza Bunge), which is used in Traditional Chinese Medicine to treat cardiovascular and other diseases, and they synthesize and accumulate in the root periderm of S. miltiorrhiza. However, there is no relevant report on the initial stage of tanshinone synthesis, as well as the root structure and gene expression characteristics. The present study aims to provide new insights into how these bioactive principles begin to synthesize by characterizing possible differences in their biosynthesis and accumulation during early root development from both spatial and temporal aspects. The morphological characteristics and the content of tanshinones in roots of S. miltiorrhiza were investigated in detail by monitoring the seedlings within 65 days after germination (DAGs). The ONT transcriptome sequencing was applied to investigate gene expression patterns. The periderm of the S. miltiorrhiza storage taproot initially synthesized tanshinone on about 30 DAGs. Three critical stages of tanshinone synthesis were preliminarily determined: preparation, the initial synthesis, and the continuous rapid synthesis. The difference of taproots in the first two stages was the smallest, and the differentially expressed genes (DEGs) were mainly enriched in terpene synthesis. Most genes involved in tanshinone synthesis were up regulated during the gradual formation of the red taproot. Plant hormone signal transduction and ABC transport pathways were widely involved in S. miltiorrhiza taproot development. Five candidate genes that may participate in or regulate tanshinone synthesis were screened according to the co-expression pattern. Moreover, photosynthetic ferredoxin (FD), cytochrome P450 reductase (CPR), and CCAAT binding transcription factor (CBF) were predicted to interact with the known downstream essential enzyme genes directly. The above results provide a necessary basis for analyzing the initial synthesis and regulation mechanism of Tanshinones.