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. 2021 Jun 14:12:683037.
doi: 10.3389/fgene.2021.683037. eCollection 2021.

De novo Transcriptome Sequencing Coupled With Co-expression Analysis Reveal the Transcriptional Regulation of Key Genes Involved in the Formation of Active Ingredients in Peucedanum praeruptorum Dunn Under Bolting Period

Cheng Song  1   2 , Xiaoli Li  2   3 , Bin Jia  2   3 , Li Liu  2   3 , Jinmei Ou  3 , Bangxing Han  1   2   3
Affiliations

De novo Transcriptome Sequencing Coupled With Co-expression Analysis Reveal the Transcriptional Regulation of Key Genes Involved in the Formation of Active Ingredients in Peucedanum praeruptorum Dunn Under Bolting Period

Cheng Song et al. Front Genet. .

Abstract

Peucedanum praeruptorum Dunn is a perennial and one-off flowering plant of the Peucedanum genus in Umbelliferae. The cultivated P. praeruptorum Dunn usually grows nutritionally in the first year and then moves into the reproductive growth in the second year. The lignification of the roots caused by bolting leads to the quality decline of crude materials. Since most of the previous studies have dealt with coumarin biosynthesis and identification of functional genes in P. praeruptorum, the scientific connotation of the inability that the bolted P. praeruptorum cannot be used medically is still unclear. Here, we employed a transcriptome sequencing combined with coexpression analysis to unearth the regulation mechanism of key genes related to coumarin synthesis in pre- and postbolting period, and to explore the mechanisms underlying the effects of bolting on the formation and transport of coumarins between the annual and biennial plants. Six cDNA libraries were constructed, and the transcripts were sequenced and assembled by Illumina Hiseq platform. A total of 336,505 unigenes were obtained from 824,129 non-redundant spliced transcripts. Unigenes (114,488) were annotated to the NCBI nr database, 119,017 and 10,475 unigenes were aligned to Gene Ontology (GO) functional groups and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways, respectively. Differential expression analysis screened out a series of upregulated and downregulated genes related to the phenylpropanoid pathway. The heatmap clustering showed that the similar expression patterns were both observed in groups C vs. D and groups C vs. F. The WGCNA-based coexpression was performed to elucidate the module and trait relationship to unearth important genes related to the bolting process. Seven pivotal modules on the KEGG functional annotations suggested these genes were mainly enriched in the process of plant-pathogen interaction, plant hormone signal transduction, MAPK signaling pathway, α-linolenic acid metabolism, circadian rhythm, and phenylpropanoid pathway. Further analysis provided clues that the key genes of the phenylpropanoid pathway, the ABC transporters, the apoptosis-related and circadian rhythm regulatory genes may play pivotal roles in regulating bolting signaling, biosynthesis, and transportation of coumarins.

Keywords: Peucedanum praeruptorum; bolting period; coexpression analysis; coumarins; key gene; transcriptional regulation.

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Conflict of interest statement

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest. The handling editor declared a past collaboration with one of the authors CS.

Figures

FIGURE 1
FIGURE 1
The assembly and sequence length distribution of transcripts and unigenes. (A) The GC content distribution of the transcripts. (B) The GC content distribution of the unigenes. (C) The sequence length distribution of the transcripts. (D) The sequence length distribution of unigenes. (E) The isoform number of unigenes upon alternative splicing.
FIGURE 2
FIGURE 2
The gene function annotation and the CDS prediction. (A) The ratio line chart of the main database annotation. (B) The Venn map of the annotated genes based on NR, Kyoto Encyclopedia of Genes and Genomes (KEGG), Swissprot, and KOG. (C) The distribution of homologous species based on NR. (D) The classification of annotation function based on KOG. (E) The Gene Ontology (GO) annotation distribution of different categories. (F) The KEGG pathway classification of the annotated genes. (G) The length distribution of CDS. (H) Distribution of CDS length ratio.
FIGURE 3
FIGURE 3
The gene structure and the SNP analysis. (A) The density distribution of the SSR marker. The number on the x-axis represents the SSR repeat types. (B) The amount of SNP and InDel in different groups. (C) The SNP mutation of sample A1.
FIGURE 4
FIGURE 4
The gene expression level and the intergroup correlation analysis. (A) The TPM density distribution between groups. (B) Heatmap cluster analysis of the correlation among samples. (C) The Venn of coexpression and specific genes. (D) PCA visualization of the different groups. (E) PCoA visualization of the different groups.
FIGURE 5
FIGURE 5
The differential gene expression analysis of P. praeruptorum. (A) The amount of up- and downregulated DEGs in different groups. (B) Scatter plots of the DEGs in A vs. B, A vs. C, C vs. D, and E vs. F. (C) Heatmap of the differential genes between groups based on fold change values. The red denotes upregulated expression, and the green denotes downregulated expression.
FIGURE 6
FIGURE 6
The heatmap clustering of the differential genes between groups.
FIGURE 7
FIGURE 7
The cluster dendrogram of coexpressed genes.
FIGURE 8
FIGURE 8
The eigengene adjacency trees of different conditions. (A) Annually grown at drawn phase. (B) Annually grown at undrawn phase. (C) Annually grown in north slope. (D) Annually grown in south slope. (E) Biennially grown at drawn phase. (F) Biennially grown at undrawn phase.
FIGURE 9
FIGURE 9
The module–trait relationship between six conditions and modules.
FIGURE 10
FIGURE 10
The scatter plots of the significant KEGG enrichment of functional pathway. (A) The highly enriched pathways of the black module. (B) The highly enriched pathways of the blue module. (C) The highly enriched pathways of the brown module. (D) The highly enriched pathways of the green module. (E) The highly enriched pathways of the red module. (F) The highly enriched pathways of the turquoise module. (G) The highly enriched pathways of the yellow module. The vertical axis is the function annotation information, and the horizontal axis is rich factor (equal to the number of differential genes annotated to the function divided by the number of genes annotated to the function). Q value is shown in different colors, and the number of different genes is shown in the size of dots.
FIGURE 11
FIGURE 11
A plausible model on the molecular mechanism of photoperiod regulating bolting and transformation of coumarins in P. praeruptorum. PHYA (TRINITY_DN77328_c0_g1), PIF3 (TRINITY_DN83313_c0_g2), PAP1 (TRINITY_DN63777_c0_g1, TRINITY_DN63757_c0_g2), CRY1 (TRINITY_DN82168_c0_g4, TRINITY_DN96996_c3_g1), GI (TRINITY_DN87544_c1_g2), CO (TRINITY_DN75433_c0_g3, TRINITY_DN94065_c2_g1), FT (TRINITY_DN110183_c0_g1, TRINITY_DN61953_c0_g1), HY5 (TRINITY_DN89278_c0_g1, TRINITY_DN5919_c0_g1), CHS (TRINITY_DN70261_c1_g1, TRINITY_DN91517_c0_g1, TRINITY_DN80066_c0_g1), CytC (TRINITY_DN69779_c1_g2, TRINITY_DN70114_c2_g1).
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