Enhanced Synthesis of Volatile Compounds by UV-B Irradiation in Artemisia argyi Leaves

Abstract

Background: Volatile compounds have a deep influence on the quality and application of the medicinal herb Artemisia argyi; however, little is known about the effect of UV-B radiation on volatile metabolites. Methods: We herein investigated the effects of UV-B exposure on the volatile compounds and transcriptome of A. argyi to assess the potential for improving its quality and medicinal characteristics. Results: Out of 733 volatiles obtained, a total of 133 differentially expressed metabolites (DEMs) were identified by metabolome analysis. These were classified into 16 categories, primarily consisting of terpenoids, esters, heterocyclic compounds, alcohols, and ketones. Sensory odor analysis indicated that green was the odor with the highest number of annotations. Among the 544 differentially expressed genes (DEGs) identified by transcriptome analysis, most DEGs were linked to "metabolic pathways" and "biosynthesis of secondary metabolites". Integrated analysis revealed that volatiles were mainly synthesized through the shikimate pathway and the MEP pathway. RNA-seq and qPCR results indicated that transcription factors HY5, bHLH25, bHLH18, bHLH148, MYB114, MYB12, and MYB111 were upregulated significantly after UV-B radiation, and were therefore considered key regulatory factors for volatiles synthesis under UV-B radiation. Conclusions: These findings not only provide new insights into UV-induced changes in volatile compounds, but also provide an exciting opportunity to enhance medicinal herbs' value, facilitating the development of products with higher levels of essential oils, flavor, and bioactivity.

Keywords: Artemisia argyi; UV-B radiation; transcription factor; volatile compound.

Figure 1

Overview of volatile metabolite changes in A. argyi leaves in response to UV-B radiation. (A) PCA of metabolites. (B) Cluster heatmap of all metabolite contents. The horizontal axis represents the sample name, and the vertical axis represents the metabolite information. Different colors are filled with different values obtained after standardizing the relative content (red represents high content, green represents low content). (C) Volcano plot of DEMs. (D) KEGG enrichment analysis of DEMs. (E) Top 20 upregulated DEMs.

Figure 2

Odorous compounds analysis. (A) Category and number of DEMs with sensory flavor. (B) Radar chart of sensory flavor characteristics of differential volatile compounds. (C) Correlation network diagram between sensory flavor characteristics and DEMs.

Figure 3

Overview of transcriptome analysis of A. argyi responsive to UV-B irradiation. (A) PCA analysis of samples taken at 0 h, 4 h, 8 h, and 6 days. (B) Changes in the total number of genes and DEGs. (C) Volcano map of DEGs from the pairwise comparison of UV0 vs. UV6d. (D) Venn graph for up- and downregulated DEGs from the pairwise comparisons of UV0 vs. UV4h, UV0 vs. UV8h, and UV0 vs. UV6d.

Figure 4

Enrichment analysis of metabolic pathways in comparison of UV0 vs. UV6d. (A) Top 20 enriched GO pathways of DEGs. (B) Top 20 enriched KEGG pathways of DEGs. The color and size of the solid circles represent the significant value of the enrichment factor and the number of transcripts involved in the specific pathway, respectively. (C) Classification of enriched metabolic pathways. The numbers in the figure represent the number of DEGs annotated to this pathway, and the parentheses indicate the ratio of DEGs annotated to this pathway to the number of background genes annotated to this pathway.

Figure 5

Metabolic analysis of volatile compounds in A. argyi leaves. HK, hexokinase; PFK, 6-phosphofructokinase; PK, pyruvate kinase; MEP, 2-C-methyl-D-erythrin-4-phosphate; IPP, isopentenyl pyrophosphate; DXS, 1-Deoxy-D-xylulose-5-phosphate synthase; DXR, 1-deoxy-d-xylulose-5-phosphate reductoisomerase; CMS, 2-C-methyl-D-erythritol 4-phosphate cytidine synthase; CMK, 2-C-Methyl-D-erythritol 4-phosphate cytidine kinase; MCS, 2-C-methyl-D-erythritol 2,4-cyclodiphosphate synthase; HDS, Hydroxymethylbutene-4-phosphate synthase; HDR, 1-Hydroxy-2-methyl-2 (E)-butenyl-4-diphosphate reductase; GGPPS, geranylgeranyl diphosphate synthases; FPPS, Farnesyl pyrophosphate synthase; Cit2, citrate synthase; Icl, isocitrate lyase; Idh, isocitrate dehydrogenase; Kgd, alpha-ketoglutarate dehydrogenase; Sdh, succinate dehydrogenase; Fum, fumarase; Mdh, malate dehydrogenase; DS, DAHP synthase; DAS, 3-dehydroquinic acid synthase; DAD, 3-dehydroquinic acid dehydratase; SDH, shikimate dehydrogenase; SK, shikimate kinase; ES, EPSP synthase; BAS, branched acid synthase; ICS, isochorismate synthase; PBS3, avrPphB susceptible 3; PAL, phenylalanine ammonia-lyase; C4H, cinnamate4-hydroxylase; 4CL, 4-coumarate-CoA ligase. The results were expressed as mean ± SD of triplicate measurements.

Figure 6

Transcriptional regulation of volatile compounds induced by UV-B. Gene expression of transcription factors analyzed by RNA-seq (A) and qPCR (B). Red characters indicate the upregulated metabolites. (C) A regulation model of volatile compounds-related genes. Red characters indicate the upregulated genes. Black characters indicate expression with insignificant differences. UVR8, UV Resistance Locus 8; COP1, Constitutively Photomorphogenetic 1; HY5, Elongated Hypocotyl 5.