Transcriptomic Responses of Cordyceps militaris to Salt Treatment During Cordycepins Production

Abstract

Cordycepin is a major bioactive compound found in Cordyceps militaris (C. militaris) that exhibits a broad spectrum of biological activities. Hence, it is potentially a bioactive ingredient of pharmaceutical and cosmetic products. However, overexploitation and low productivity of natural C. militaris is a barrier to commercialization, which leads to insufficient supply to meet its existing market demands. In this study, a preliminary study of distinct concentrations of salt treatments toward C. militaris was conducted. Although the growth of C. militaris was inhibited by different salt treatments, the cordycepin production increased significantly accompanied by the increment of salt concentration. Among them, the content of cordycepin in the 7% salt-treated group was five-fold higher than that of the control group. Further transcriptome analysis of samples with four salt concentrations, coupled with Gene Ontology (GO) analysis and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment, several differentially expressed genes (DEGs) were found. Finally, dynamic changes of the expression patterns of four genes involved in the cordycepin biosynthesis pathway were observed by the quantitative real-time PCR. Taken together, our study provides a global transcriptome characterization of the salt treatment adaptation process in C. militaris and facilitates the construction of industrial strains with a high cordycepin production and salt tolerance.

Keywords: Cordyceps militaris; cordycepin; molecular mechanism; salt treatment; transcriptome.

Figure 1

The phenotype of C. militaris is affected by different salt treatments. 0, 3, 5, and 7% are equivalent to the control, slight, moderate, and severe salt treatment, respectively.

Figure 2

The contents of cordycepin with mg/g dry weight 0, 3, 5, and 7% are equivalent to the control, slight, moderate, and severe salt treatment, respectively. Asterisks indicate statistically significant differences between groups (Student's t-test): * and ** indicating significance level was accepted at P ≤ 0.05 and P < 0.01, respectively, as compared to control.

Figure 3

Distribution of differentially expressed genes (DEGs) in the six samples. (A) DEGs' distribution between control and three different salt treatments; (B) Venn diagram of the commonly expressed genes between samples. NaCl-3, 5, and 7 represent 3, 5, and 7% salt concentrations, respectively.

Figure 4

GO classification and enrichment of the DEGs based on the GO database [the x-axis represents the GO term and enrichment, and the y-axis represents the percentage of genes (left) and the number of DEGs (right)]. Selected_gene_number represents the number of DEGs. GO, gene oncology; DEGs, differentially expressed genes.

Figure 5

The functional enrichment analysis of DEGs using KEGG annotation. The x-axis represents the rich factor, and the y-axis represents the pathway name. DEGs, differentially expressed genes; KEGG, kyoto encyclopedia of genes and genomes.

Figure 6

Cordycepin biosynthesis pathway in C. militaris and qRT-PCR results. Bar chart with 4 colors represents the relative expression levels of four genes in response to distinct salt treatments. The bars represent the average (±SE) of biological repeats. NaCl-3, 5, and 7 represent 3%, 5%, and 7% salt concentrations, respectively. Asterisks indicate statistically significant differences between groups (Student's t-test): * and ** indicating significance level was accepted at P ≤ 0.05 and P < 0.01, respectively, ns: no significant difference, as compared to control.