Comparative Metabolome and Transcriptome Analyses of the Regulatory Mechanism of Light Intensity in the Synthesis of Endogenous Hormones and Anthocyanins in Anoectochilus roxburghii (Wall.) Lindl

Abstract

To explore the regulatory mechanism of endogenous hormones in the synthesis of anthocyanins in Anoectochilus roxburghii (Wall.) Lindl (A. roxburghii) under different light intensities, this study used metabolomics and transcriptomics techniques to identify the key genes and transcription factors involved in anthocyanin biosynthesis. We also analyzed the changes in and correlations between plant endogenous hormones and anthocyanin metabolites under different light intensities. The results indicate that light intensity significantly affects the levels of anthocyanin glycosides and endogenous hormones in leaves. A total of 38 anthocyanin-related differential metabolites were identified. Under 75% light transmittance (T3 treatment), the leaves exhibited the highest anthocyanin content and differentially expressed genes such as chalcone synthase (CHS), flavonol synthase (FLS), and flavonoid 3'-monooxygenase (F3'H) exhibited the highest expression levels. Additionally, 13 transcription factors were found to have regulatory relationships with 7 enzyme genes, with 11 possessing cis-elements responsive to plant hormones. The expression of six genes and two transcription factors was validated using qRT-PCR, with the results agreeing with those obtained using RNA sequencing. This study revealed that by modulating endogenous hormones and transcription factors, light intensity plays a pivotal role in regulating anthocyanin glycoside synthesis in A. roxburghii leaves. These findings provide insights into the molecular mechanisms underlying light-induced changes in leaf coloration and contribute to our knowledge of plant secondary metabolite regulation caused by environmental factors.

Keywords: RNA-seq; correlation analysis; phytohormone; promoter cis-acting element; qRT-PCR validation; transcription factor.

Figure 1

Heat map of the clustering of differential anthocyanin glycoside metabolites in A. roxburghii (Wall.) Lindl leaves at different light intensities.

Figure 2

Heat map of clustering of different phytohormone metabolites in A. roxburghii leaves under different light intensities.

Figure 3

Heat map of clustering of differential genes related to anthocyanins glycoside synthesis in A. roxburghii leaves at different light intensities.

Figure 4

Conjoint analysis of transcriptome metabolism of anthocyanins in A. roxburghii leaves under different light intensities.

Figure 5

Correlation network of “phytohormone–anthocyanin glycoside genes” in A. roxburghii leaves at different light intensities. The solid line indicates positive regulation, the dashed line indicates negative regulation, and the thicker the line, the stronger the correlation of regulation.

Figure 6

Correlation network of “phytohormone–anthocyanidin glycoside–related metabolites” in A. roxburghii leaves at different light intensities. The solid line indicates positive regulation, the dashed line indicates negative regulation, and the thicker the line, the stronger the correlation of regulation.

Figure 7

Regulatory network diagram of “transcription factor–anthocyanin metabolism-related enzyme gene expression” in A. roxburghii leaves under different light intensities. The solid line indicates positive regulation, the dashed line indicates negative regulation, and the thicker the line, the stronger the correlation of regulation.

Figure 8

Analysis of the promoter cis-elements of anthocyanin-related transcription factors in the leaves of A. roxburghii under different light intensities.

Figure 9

qRT-PCR analysis of differential enzymes and transcription factors in A. roxburghii leaves under different light intensity treatments.