Drought Stress Stimulates the Terpenoid Backbone and Triterpenoid Biosynthesis Pathway to Promote the Synthesis of Saikosaponin in Bupleurum chinense DC. Roots

Abstract

Bupleurum chinense is an important medicinal plant in China; however, little is known regarding how this plant transcribes and synthesizes saikosaponins under drought stress. Herein, we investigated how drought stress stimulates the transcriptional changes of B. chinense to synthesize saikosaponins. Short-term drought stress induced the accumulation of saikosaponins, especially from the first re-watering stage (RD_1 stage) to the second re-watering stage (RD_2 stage). Saikosaponin-a and saikosaponin-d increased by 84.60% and 75.13%, respectively, from the RD_1 stage to the RD_2 stage. Drought stress also stimulated a rapid increase in the levels of the hormones abscisic acid, salicylic acid, and jasmonic acid. We screened 49 Unigenes regarding the terpenoid backbone and triterpenoid biosynthesis, of which 33 differential genes were significantly up-regulated during drought stress. Moreover, one P450 and two UGTs are possibly involved in the synthesis of saikosaponins, while some transcription factors may be involved in regulating the expression of key enzyme genes. Our study provides a reference for the cultivation of B. chinense and a practical means to ensure the quality (safety and effectiveness) of B. chinense for medicinal use, as well as insights into the modernization of the China Agriculture Research System.

Keywords: Bupleurum chinense DC.; drought stress; plant hormones; saikosaponins; terpenoid backbone and triterpenoid biosynthesis.

Figure 1

Changes in saikosaponin content under drought stress. (A) Changes in saikosaponin content. (B) Changes in SS-a content. (C) Changes in SS-d content. (D) Changes in SS-c content. (E) Changes in SS-e content. (F) Changes in SS-f content. Data are expressed as means ± SDs (n = 3). * indicates that the drought group and control group significantly differed at the 0.05 level.

Figure 2

Plant hormone content in roots of B. chinense under drought stress. (A) Changes in the abscisic acid content. (B) Changes in the salicylic acid content. (C) Changes in the jasmonic acid content. Data are expressed as means ± SDs (n = 3). * indicates that the drought group and control group significantly differed at the 0.05 level.

Figure 3

Principal component analysis (PCA) biplot of the first two PCA axes for different B. chinense samples under drought stress. Data are expressed as means (n = 3).

Figure 4

Analysis of differentially expressed genes in B. chinense samples under drought stress based on transcriptome sequencing. (A) Up-regulation and down-regulation of differentially expressed genes. (B) Venn diagrams of differentially expressed genes among different samples. (C) PCA of different samples. (D) Cluster analysis of differentially expressed genes.

Figure 5

KEGG enrichment of the terpenoid backbone and triterpenoid biosynthesis (A) and cluster analysis of the terpenoid backbone and triterpenoid biosynthesis differentially expressed genes (B).

Figure 6

Differentially expressed genes annotated to P450 (A) and UGT (B) and their cluster analysis.

Figure 7

Differentially expressed genes annotated to ERF (A), BHLB (B), MYB (C), WRKY (D), and NAC (E) and their cluster analysis.

Figure 8

Effects of drought stress on the expression of the key saikosaponin biosynthesis enzyme genes. Expression levels of (A) HMGR gene, (B) IPPI gene, (C) FPS gene, (D) SS gene, (E) SE gene, and (F) β-AS gene. Data are expressed as means ± SDs (n = 3). * indicates that the drought group and control group significantly differed at the 0.05 level.

Figure 9

The saikosaponin synthesis regulation network under drought stress.