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. 2022 Jun 29:10:e13670.
doi: 10.7717/peerj.13670. eCollection 2022.

Integrated transcriptome and metabolome analyses revealed regulatory mechanisms of flavonoid biosynthesis in Radix Ardisia

Chang Liu  1 , Jie Pan  1 , Zhi-Gang Yin  2 , Tingting Feng  1 , Jiehong Zhao  1 , Xiu Dong  3 , Ying Zhou  1   2
Affiliations

Integrated transcriptome and metabolome analyses revealed regulatory mechanisms of flavonoid biosynthesis in Radix Ardisia

Chang Liu et al. PeerJ. .

Abstract

Background: Radix Ardisia (Jab Bik Lik Jib) is a common Miao medicine and is widely distributed in the Guizhou region of southern China. The botanical origin of Radix Ardisia includes the dry root and rhizome of Ardisia Crenata Sims (ACS) or Ardisia Crispa (Thunb.) A.DC. (AC), which are closely related species morphologically. However, the secondary metabolites in their roots are different from one another, especially the flavonoids, and these differences have not been thoroughly explored at the molecular level. This project preliminarily identified regulatory molecular mechanisms in the biosynthetic pathways of the flavonoids between ACS and AC using a multi-omics association analysis.

Methods: In this study, we determined the total levels of saponin, flavonoid, and phenolic in Radix Ardisia from different origins. Integrated transcriptome and metabolome analyses were used to identify the differentially expressed genes (DEGs) and differentially expressed metabolites (DEM). We also performed conjoint analyses on DEGs and DEMs to ascertain the degree pathways, and explore the regulation of flavonoid biosynthesis.

Results: The total flavonoid and phenolic levels in ACS were significantly higher than in AC (P < 0.05). There were 17,685 DEGs between ACS vs. AC, 8,854 were upregulated and 8,831 were downregulated. Based on this, we continued to study the gene changes in the flavonoid biosynthesis pathway, and 100 DEGs involving flavonoid biosynthesis were differentially expressed in ACS and AC. We validated the accuracy of the RNA-seq data using qRT-PCR. Metabolomic analyses showed that 11 metabolites were involved in flavonoid biosynthesis including: Naringenin, Luteolin, Catechin, and Quercetin. A conjoint analysis of the genome-wide connection network revealed the differences in the types and levels of flavonoid compounds between ACS and AC. The correlation analysis showed that Naringenin, Luteolin, Catechin, and Quercetin were more likely to be key compounds in the flavonoid biosynthesis pathway also including 4CL, AOMT, CHS, CHI, DFR, F3'5'H, FLS, and LAR.

Conclusions: This study provides useful information for revealing the regulation of flavonoid biosynthesis and the regulatory relationship between metabolites and genes in the flavonoid biosynthesis pathway in Radix Ardisia from different origins.

Keywords: Ardisia crenata Sims; Ardisia crispa (Thunb.) A.DC.; Conjoint analysis; Flavonoid biosynthesis; Metabolomics; Transcriptomics.

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Conflict of interest statement

Xiu Dong is employed by Guizhou Sanli Pharmaceutical Co., Ltd.

Figures

Figure 1
Figure 1. Total saponin, flavonoid, and phenolic content in Radix Ardisia from different origins (mg/g).
ACS, Ardisia Crenata Sims; ACSV, Ardisia Crenata Sims var. bicolor (Walker) C. Y. Wu et C. Chen; AC, Ardisia Crispa (Thunb.) A.DC.
Figure 2
Figure 2. The differentially expressed genes between ACS and AC (A) Volcano plot; (B) MA plot.
Red dots represent upregulated DEGs and green dots represent downregulated DEGs.
Figure 3
Figure 3. The qRT-PCR analysis of the genes.
(A) The RNA-seq results revealed differentially expressed genes. (B) Differentially expressed genes were confirmed by qRT-PCR. Data are represented as mean values ± SD. n= 3.
Figure 4
Figure 4. The main terms of the KEGG analysis.
Figure 5
Figure 5. The differentially expressed genes of flavonoid biosynthesis in ACS and AC.
Figure 6
Figure 6. (A) Differentially expressed metabolites (ESI+); (B) differentially expressed metabolites (ESI-).
Figure 7
Figure 7. The KEGG pathways involving differentially expressed metabolites between ACS and AC.
(A) ESI+; (B) ESI-; (C) Differentially expressed metabolites in flavonoid biosynthesis.
Figure 8
Figure 8. Heat map of flavonoid-related genes and metabolites in ACS and AC.
449: Succinate; 1321: p-Hydroxycinnamaldehyde; 740: Salicylic acid (-); 2506: Sinapic acid; 1549: L-Tyrosine; 2138: N-Acetyl-L-phenylalanine; 1819: Ferulic acid; 3093: Alpha-N-Phen.
Figure 9
Figure 9. Analysis of the genome-wide connection network between genes and metabolites related to flavonoids in ACS and AC.
The purple triangle represents metabolites and the blue circle represents genes.
See this image and copyright information in PMC

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