Identification of drought-responsive phenolic compounds and their biosynthetic regulation under drought stress in Ligularia fischeri

Ligularia fischeri, a leafy edible plant found in damp shady regions, has been used as an herbal medicine and is also consumed as a horticultural crop. In this study, we investigated the physiological and transcriptomic changes, especially those involved in phenylpropanoid biosynthesis, induced by severe drought stress in L. fischeri plants. A distinguishing characteristic of L. fischeri is a color change from green to purple due to anthocyanin biosynthesis. We chromatographically isolated and identified two anthocyanins and two flavones upregulated by drought stress using liquid chromatography-mass spectrometry and nuclear magnetic resonance analyses in this plant for the first time. In contrast, all types of caffeoylquinic acids (CQAs) and flavonol contents were decreased under drought stress. Further, we performed RNA sequencing to examine the molecular changes in these phenolic compounds at the transcriptome level. In an overview of drought-inducible responses, we identified 2,105 hits for 516 distinct transcripts as drought-responsive genes. Moreover, differentially expressed genes (DEGs) associated with phenylpropanoid biosynthesis accounted for the greatest number of both up- and downregulated DEGs by Kyoto Encyclopedia of Genes and Genomes enrichment analysis. We identified 24 meaningful DEGs based on the regulation of phenylpropanoid biosynthetic genes. Potential drought-responsive genes included upregulated flavone synthase (LfFNS, TRINITY DN31661 c0 g1 i1) and anthocyanin 5-O-glucosyltransferase (LfA5GT1, TRINITY DN782 c0 g1 i1), which could contribute to the high levels of flavones and anthocyanins under drought stress in L. fischeri. In addition, the downregulated shikimate O-hydroxycinnamolytransferase (LfHCT, TRINITY DN31661 c0 g1 i1) and hydroxycinnamoyl-CoA quinate/shikimate transferase (LfHQT4, TRINITY DN15180 c0 g1 i1) genes led to a reduction in CQAs. Only one or two BLASTP hits for LfHCT were obtained for six different Asteraceae species. It is possible that the HCT gene plays a crucial role in CQAs biosynthesis in these species. These findings expand our knowledge of the response mechanisms to drought stress, particularly regarding the regulation of key phenylpropanoid biosynthetic genes in L. fischeri.