Integrative analysis of the metabolome and transcriptome provides insights into the mechanisms of flavonoid biosynthesis in Polygonatum

Abstract

A noteworthy group of culinary and medicinal plants is Polygonatum species. They are known for their abundant flavonoid compound-rich rhizomes, which have antioxidative and anticancer activities. Using Polygonatum sibiricum Red (SXHZ) and Polygonatum kingianum var. grandifolium (HBES), we conducted transcriptome and metabolomic investigations to look into the molecular processes that control the manufacture of these flavonoids in Polygonatum plants. Seven distinct flavonoid metabolites were identified by the analytical data, with phloretin exhibiting a notable differential expression in the biosynthetic pathway. 30 genes with differential expression were found in both plants after further investigation, five of which are members of the transcription factor family associated with MBW. Thus, we suggest that Phloretin and the genes belonging to the MYB-related transcription factor family play a crucial role in controlling the flavonoid biosynthesis pathway in Polygonatum. This work lays the groundwork for a deeper comprehension of the biosynthesis and metabolic processes of flavonoids in Polygonatum, serving as an invaluable resource for the development of the polygonatum-related pharmaceutical industries as well as for the future breeding of Polygonatum plants with higher flavonoid content.

Fig 1. HBES and SXHZ multivariate statistical analyses.

A: overall sample PCA plot; B: PLS-DA plot for latent structures; C: OPLS-DA validation plot; D: OPLS-DA score plot.

Fig 2. Analysis of DEMs and KEGG statistics.

A: DEMs of the overall sample. B. Metabolites in the top 20 multiplicity of differences in the comparative group. C. KEGG pathway annotations of the DEMs.

Fig 3. Map of annotation results of two Polygonatum gene databases.

A: Venn diagram of 7 major database annotations. B: KOG database annotations. C: KEGG database annotations. D: GO database annotations.

Fig 4. Co-expression profiles of all transcripts and active components in two Polygonatum rhizomes.

A: Hierarchical clustering tree of the co-expressed modules of two Polygonatum species. B: Heat map of the correlation of the two Polygonatum samples and modules. C: Network diagram depicting the top 50 ranked linkage strengths of genes in the MElightgreen module. D: Network diagram illustrating the top 50 ranked linkage strengths of genes in the MEred module. E: Network diagram displaying the top 50 ranked linkage strengths of genes in the MEyellow module. F: Network diagram showing the top 50 ranked linkage strengths of genes in the MEblack module. G: Network diagram presenting the top 50 ranked linkage strengths of genes in the MEgrey module.

Fig 5. Differential gene and differential metabolite analysis.

A: Enrichment circle plot of differentially expressed genes in the GO database for Polygonatum, the bar graph shows the proportion of up-regulated genes, and the red color represents the number of up-regulated genes. B: Number of genes in the top 20 enriched TFs family genes. C: Number of genes in the differentially expressed TFs family genes. D: Number of total genes in the KEGG pathway and the number of DEGs (the abscissa A in the middle of the pathway represents Phenylpropanoid biosynthesis; B represents Flavone and flavonol biosynthesis; C represents Flavonoid biosynthesis; D represents Isoflavonoid biosynthesis). E: FC between transcriptome and metabolome correlation heatmap of the top 20 enriched DEGs and DEGs. F: Correlation network of the top 20 DEGs and all DEMs associated with the flavonoid biosynthesis pathway. G: DEGs and DEMs involved in the flavonoid biosynthesis pathway.

Fig 6. Study of flavonoid biosynthesis and metabolic pathways in Polygonatum rhizomes.

Blue dots indicate that the metabolite was detected but not significantly changed, and green dots indicate that the metabolite content was significantly down-regulated in the experimental group. Red rectangles indicate that the enzyme is associated with up-regulated genes, green rectangles indicate that the enzyme is associated with down-regulated genes, blue rectangles indicate that the enzyme is associated with both up- and down-regulated genes, and the numbers in the boxes represent the enzyme numbers (EC numbers).

Fig 7. Results of qRT-PCR assay.

The bar graph represents the FPKM results of transcriptome data; the line graph represents the qRT-PCR results.