SmNAC68 transcription factor negatively regulate tanshinone biosynthesis under low-temperature conditions in Salvia miltiorrhiza.

Salvia miltiorrhiza Bge. (S. miltiorrhiza) is a traditional Chinese herbal medicine, primarily used for treating cardiovascular diseases. Tanshinones as the main medicinal components of S. miltiorrhiza, their content is an important factor affecting the medicinal and economic value. Reasonable post-harvest techniques are the critical means to improve the effective components, and short-term low-temperature can induce the synthesis of plant secondary metabolites. However, the molecular mechanisms of low-temperature regulation of tanshinones remain unclear. This study found that post-harvest low-temperature can significantly accumulate the tanshinone components, simultaneously, it markedly enhances the expression of key enzyme genes involved in tanshinone biosynthesis. Analysis of the NAC gene family revealed that SmNAC68 is predominantly active in roots, with a marked decrease in expression under low-temperature. Genetic transformation experiments demonstrated that the accumulation of four tanshinones in SmNAC68 overexpression individual was significantly downregulated, with the key enzyme gene SmCPS1 showing the most significant downregulation. Protein interaction experiments revealed that SmNAC68 can interact with the promoter of SmCPS1 and inhibit its transcription. The results reveal the molecular mechanism of low-temperature regulation of tanshinone biosynthesis, which is of great significance for improving the post-harvest processing and quality enhancement of S. miltiorrhiza.