Transcriptomics and network pharmacology analysis reveal key genes in alkaloid biosynthesis in Zephyranthes candida and therapeutic targets for LIHC.

Main conclusion: This study identified key enzyme-encoding genes involved in the biosynthesis of lycorine and galanthamine in Zephyranthes candida, and predicted four critical therapeutic targets of lycorine for liver hepatocellular carcinoma (LIHC). Amaryllidaceae alkaloids (AAs) comprise a unique class of isoquinoline alkaloids mainly found in plants of the Amaryllidaceae family. Zephyranthes candida (Lindl.) Herb., a bulbous geophyte within this family, produces lycorine and galanthamine as its characteristic isoquinoline alkaloids. In this study, an in-depth analysis of the biosynthesis of these compounds in various tissues of the plant was conducted using transcriptomics and bioinformatics. Transcriptome databases were generated for roots, leaves, and bulbs of Z. candida. Key enzymes and differentially expressed genes (DEGs) associated with lycorine and galanthamine biosynthesis were screened, leading to the identification of two genes, ZcTYDC (DN4967_1) and ZcN4OMT (DN3779), based on correlation analysis. Subsequently, phylogenetic, structural modeling and molecular docking analyses of key enzymes were undertaken, followed by a detailed assessment of their characteristics and functions. A correlation was identified between the expression of genes encoding transcription factors (TFs) and major AAs biosynthetic enzymes in Z. candida. The expression levels of selected genes were determined by quantitative real-time polymerase chain reaction (qPCR), while the contents of lycorine and galanthamine in each tissue were detected by high-performance liquid chromatography (HPLC). Critical targets of lycorine in liver hepatocellular carcinoma (LIHC)-MMP9, IGF1, SRC, and CCNA2-were predicted using network pharmacology, gene expression profiling interactive analysis (GEPIA2), molecular docking, and visualization. This study provides a theoretical basis for further research on the biosynthesis of AAs, as well as support for the therapeutic application of lycorine in LIHC.