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. 2022 Jun 23;17(6):e0270309.
doi: 10.1371/journal.pone.0270309. eCollection 2022.

Transcriptome analysis reveals regulation mechanism of methyl jasmonate-induced terpenes biosynthesis in Curcuma wenyujin

Qiuhui Wei  1   2 , Kaer Lan  1   2 , Yuyang Liu  1   2 , Rong Chen  2 , Tianyuan Hu  1   2 , Shujuan Zhao  1   2 , Xiaopu Yin  1   2 , Tian Xie  1   2
Affiliations

Transcriptome analysis reveals regulation mechanism of methyl jasmonate-induced terpenes biosynthesis in Curcuma wenyujin

Qiuhui Wei et al. PLoS One. .

Abstract

Curcuma wenyujin is the source plant of three traditional Chinese medicines, which have been widely used in clinical treatment over 1000 years. The content of terpenes, the major medicinal active ingredients, is relatively low in this plant. Studies have shown that MeJA can promote terpenes biosynthesis in plants. However, the mechanism underlying the effect of MeJA in C. wenyujin remains unclear. In this work, the transcriptome of C. wenyujin leaves with MeJA treatment was analyzed to elucidate the regulation mechanism of MeJA-mediated terpene biosynthesis. Based on the RNA-seq data, 7,246 unigenes were differentially expressed with MeJA treatment. Expression pattern clustering of DEGs revealed that unigenes, related to JA biosynthesis and signal transduction, responded to exogenous MeJA stimulation on the early stage and maintained throughout the process. Subsequently, unigenes related to terpene biosynthesis pathway showed a significant up-regulation with 6 h treatment. The analysis results suggested that MeJA induced the expression of JA biosynthesis genes (such as LOXs, AOSs, AOCs, OPRs, and MFPs) and JA signal transduction core genes (JAZs and MYCs) to activate JA signaling pathway. Meanwhile, downstream JA-responsive genes presented up-regulated expression levels such as AACT, HMGSs, HMGRs, DXSs, DXRs, MCTs, HDSs, and HDRs, thus promoting terpenes biosynthesis. The transcriptional expressions of these genes were validated by qRT-PCR. In addition, six CwTPS genes in response to MeJA were identified. With MeJA treatment, the expression levels of CwTPSs were increased as well as those of the transcription factors MYB, NAC, bZIP, WRKY, AP2/ERF, and HLH. These TFs might potentially regulate terpenes biosynthesis. These results provide insights for regulation mechanism of terpenes biosynthesis.

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Conflict of interest statement

The authors have declared that no competing interests exist.

Figures

Fig 1
Fig 1. Comparative statistics of differentially expressed genes (DEGs) in MeJA-treated C. wenyujin.
(A) Total number of up-regulated and down regulated genes in different groups (group 1: control vs 1 h; group 2: control vs 6 h). (B-D) Venn diagram illustrating the shared and unique DEGs between different groups. The overlaps indicate the number of shared DEGs between different groups.
Fig 2
Fig 2. Patterns of gene expression across three time points in C. wenyujin with MeJA treatment.
(A) Expression trend clustering of all DEGs. Each rectangle represents a profile. Profiles with the same color can be grouped into the same cluster. The colorless rectangle indicates that clustering is not significant. The number in the upper left corner of the rectangle is the number of profile, in which the broken line is the trend of expression over time. The p-value in the lower left corner reflects the corresponding significance level. (B-F) Expression patterns of each significant clustering profile. The number of unigenes in each profile is labeled above the frame. Broken lines represent the trend of gene expression. The numbers on the x-axis indicate the time points of 0, 1, and 6 h, respectively. The numbers on the y-axis represent up-regulation (> 0), no change (= 0), and down-regulation (< 0), respectively.
Fig 3
Fig 3
GO enrichment analysis of DEGs in C. wenyujin with MeJA-treatment for “control vs 1 h” (A) and “control vs 6 h” (B) groups. The y-axis on the right side represents the GO terms and x-axis represents rich factor. The size of the dot indicates the number of genes in this GO term, and the color of the dot corresponds to different FDR (p-vaule). p-vaule < 0.05 indicates significant enrichment of GO function. Top 20 of significant enrichment results are displayed.
Fig 4
Fig 4
KEGG enrichment analysis of DEGs in C. wenyujin with MeJA-treatment for “control vs 1 h” (A) and “control vs 6 h” (B) groups. The y-axis on the right side represents the KEGG pathway and x-axis represents rich factor. The size of the dot indicates the number of genes in KEGG pathway, and the color of the dot corresponds to different FDR (p-vaule). p-vaule < 0.05 indicates significant enrichment of KEGG pathway. Up to top 30 of significant enrichment results are displayed.
Fig 5
Fig 5. Analysis of DEGs related to terpene backbone biosynthesis pathway in C. wenyujin with MeJA-treatment.
(A) Gas chromatogram of leaves extracts from plant sprayed with or without MeJA. (B) Contents of β-elemene in leaves. (C) The terpene backbone biosynthesis pathway. Genes with red box represent enzyme genes in this biosynthesis pathway (D) Transcriptional profiles of DEGs related to terpene backbone biosynthesis pathway. The log2 (TPM) values for the DEGs were calculated based on three biological replicates for each time-point. The color scale from blue to red represents increase expression level. (E) Expression level of DEGs determined by qRT-PCR. Three independent biological replicates were performed. Asterisks indicate significant difference (*p< 0.05; **p < 0.01).
Fig 6
Fig 6. Analysis of DEGs related to JA biosynthesis and signal transduction in C. wenyujin with MeJA-treatment.
(A) JA biosynthesis and signal transduction pathway. Genes with red box represent enzyme genes in this pathway (B) Transcriptional profiles of DEGs related to JA biosynthesis and signal transduction pathway. The log2 (TPM) values for the DEGs were calculated based on three biological replicates for each time-point. The color scale from blue to red represents increase expression level. (C) Expression level of DEGs determined by qRT-PCR. Three independent biological replicates were performed. Asterisks indicate significant difference (*P < 0.05; **P < 0.01).
Fig 7
Fig 7. Analysis of CwTPSs induced by MeJA in C. wenyujin.
(A) Transcriptional profiles of identified CwTPSs. The log2 (TPM) values for the DEGs were calculated based on three biological replicates for each time-point. The color scale from blue to red represents increase expression level. (B) Phylogenetic tree of CwTPSs with other TPSs from different plants. (C) Expression level of randomly selected CwTPSs determined by qRT-PCR. Three independent biological replicates were performed. Asterisks indicate significant difference (*p < 0.05; **p < 0.01).
Fig 8
Fig 8. Co-expression network of differentially expressed CwTPSs and TF genes in C. wenyujin with MeJA-treatment.
Each node represents a gene with its serial number and annotation information. Red nodes are CwTPSs, and blue nodes are TFs. Connecting lines represent co-expression relationships.
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