Understanding the molecular mechanisms underlying the effects of light intensity on flavonoid production by RNA-seq analysis in Epimedium pseudowushanense B.L.Guo

Epimedium pseudowushanense B.L.Guo, a light-demanding shade herb, is used in traditional medicine to increase libido and strengthen muscles and bones. The recognition of the health benefits of Epimedium has increased its market demand. However, its resource recycling rate is low and environmentally dependent. Furthermore, its natural sources are endangered, further increasing prices. Commercial culture can address resource constraints of it.Understanding the effects of environmental factors on the production of its active components would improve the technology for cultivation and germplasm conservation. Here, we studied the effects of light intensities on the flavonoid production and revealed the molecular mechanism using RNA-seq analysis. Plants were exposed to five levels of light intensity through the periods of germination to flowering, the flavonoid contents were measured using HPLC. Quantification of epimedin A, epimedin B, epimedin C, and icariin showed that the flavonoid contents varied with different light intensity levels. And the largest amount of epimedin C was produced at light intensity level 4 (I4). Next, the leaves under the treatment of three light intensity levels ("L", "M" and "H") with the largest differences in the flavonoid content, were subjected to RNA-seq analysis. Transcriptome reconstruction identified 43,657 unigenes. All unigene sequences were annotated by searching against the Nr, Gene Ontology, and Kyoto Encyclopedia of Genes and Genomes (KEGG) databases. In total, 4008, 5260, and 3591 significant differentially expressed genes (DEGs) were identified between the groups L vs. M, M vs. H and L vs. H. Particularly, twenty-one full-length genes involved in flavonoid biosynthesis were identified. The expression levels of the flavonol synthase, chalcone synthase genes were strongly associated with light-induced flavonoid abundance with the highest expression levels found in the H group. Furthermore, 65 transcription factors, including 31 FAR1, 17 MYB-related, 12 bHLH, and 5 WRKY, were differentially expressed after light induction. Finally, a model was proposed to explain the light-induced flavonoid production. This study provided valuable information to improve cultivation practices and produced the first comprehensive resource for E. pseudowushanense transcriptomes.