Metabolic Response of Sanghuangporus baumii to Zn2+ Induction and Biosynthesis of a Key Pharmacological Component: Triterpenoid

Abstract

Triterpenoids derived from Sanghuangporus baumii exhibit potent antitumor activity, but their yields under natural conditions are relatively low due to their status as secondary metabolites. In this study, we investigated the effects of Zn²⁺ induction on the growth and triterpenoid biosynthesis of S. baumii. The results showed that 0.5 mM Zn²⁺ significantly enhanced the mycelial growth rate (0.43 ± 0.004 cm/d) and biomass (4.8 ± 0.024 g/L), representing increases of 8.71% and 16.95%, respectively, compared with the Zn0 group. This result was mainly caused by an increase in the soluble sugar content. Furthermore, 5 mM Zn²⁺ induced upregulation of genes in the mevalonate (MVA) pathway, thereby promoting triterpenoid accumulation by 167.86% compared with the Zn0 group. Transcriptome analysis identified SbHMGS as the key gene involved in triterpenoid biosynthesis under Zn²⁺ induction. Heterologous expression of SbHMGS in Saccharomyces cerevisiae confirmed its critical role in triterpenoid production. The triterpenoid (squalene) content of the engineered strain (Sc-HMGS) reached 0.88 mg/g under Zn²⁺ induction, which was 208.6% higher than in the non-induced control strain (Sc-NTC). These findings provide a foundation for optimizing the industrial fermentation condition of S. baumii and S. cerevisiae to enhance triterpenoid yields.

Keywords: HMGS gene; Saccharomyces cerevisiae; Sanghuangporus baumii; Zn; heterologous biosynthesis; triterpenoid.

Figure 1

Effects of varying Zn²⁺ concentrations on S. baumii mycelia growth and triterpenoid production: (A) colony morphology; (B) mycelial growth rates; (C) biomass; (D) total triterpenoid content; and (E) intracellular Zn²⁺ content in S. baumii mycelia. Error bars represent the standard deviation (SD) of three replicates (n = 3). Letters indicate a significant difference (p < 0.05).

Figure 2

RNA-seq analysis of S. baumii under Zn²⁺ treatment. (A) RNA gel electrophoresis of nine samples. (B) Venn diagram showing functional annotation results in six public protein databases. (C) PCA of transcript profiles from Zn0, Zn0.5, and Zn5 groups. (D) Clustering heatmap illustrating expression patterns among samples (red = upregulation; green = downregulation).

Figure 3

Analysis of DEGs in S. baumii under Zn²⁺ treatment. (A) Volcano plot of up- and downregulated DEGs from pairwise comparisons. (B) Expression patterns of DEGs s under Zn²⁺ treatment. Gray lines denote individual genes, and blue lines indicate the overall expression trend for each cluster. The number above each panel denotes the genes included in that expression pattern.

Figure 4

GO and KEGG enrichment analyses of DEGs under Zn^²⁺ treatment. (A) GO classification of DEGs between Zn0.5 and Zn0 groups. (B) GO classification of DEGs between Zn5 and Zn0 groups. (C) KEGG enrichment analysis of DEGs between Zn0.5 and Zn0 groups. (D) KEGG enrichment analysis of DEGs between Zn5 and Zn0 groups. (E) KEGG enrichment analysis of DEGs between Zn5 and Zn0.5 groups.

Figure 5

Changes in sugar metabolism of S. baumii mycelia induced by Zn²⁺. (A) Soluble sugar content in S. baumii under Zn²⁺ induction. (B) DEGs involved in starch and sucrose metabolism. Error bars represent the standard deviation (SD) of three replicates (n = 3). Letters indicate a significant difference (p < 0.05).

Figure 6

DEGs involved in terpenoid backbone biosynthesis. (A) Proposed pathways for triterpenoid biosynthesis in S. baumii. (B) Fold change of each MVA pathway gene in Zn5 vs. Zn0, where the numbers above the bars indicate the fold change. (C) Correlation analysis among the triterpenoid content, Zn²⁺ content, and gene expression level.

Figure 7

Heterologous expression of SbHMGS in S. cerevisiae. (A) Schematic of the pYES2-HMGS vector construction. (B) PCR product gel electrophoresis of four positive single colonies. (C) Squalene content in Sc-NTC and Sc-HMGS under Zn²⁺ induction. Error bars represent the standard deviation (SD) of three replicates (n = 3). Letters indicate a significant difference (p < 0.05).