Integrated metabolomic and transcriptomic profiling provide insights into the biosynthetic mechanism of flavonoids in the plumule of Asian lotus (Nelumbo nucifera)

The lotus plumule is a pharmacologically important tissue rich in flavonoids; yet the molecular mechanisms underlying their biosynthesis remain poorly understood. In this study, we integrated metabolomic and transcriptomic profiling to systematically elucidate the biosynthetic and regulatory pathways during plumule development. A total of 59 flavonoid compounds were identified, with flavone glycosides being the predominant class. We observed a continuous increase in total flavonoid content from 12 to 24 days after pollination. Through transcriptome profiling, we reconstructed the flavonoid biosynthetic pathway and identified two candidate C -glycosyltransferase ( CGT ) genes. Functional validation via transient overexpression assays in lotus petals confirmed that NnCGTa promotes the accumulation of flavonoid C -glycosides, such as schaftoside and vitexin. Additionally, we characterized the highly expressed R2R3-MYB transcription factor NnMYB12 as a potential key regulatory hub through in vivo assays. Its transient overexpression significantly increased flavonoid accumulation and upregulated the expression of flavonoid biosynthetic structural genes in lotus petals and tobacco leaves. In vivo experiments demonstrated that NnMYB12 directly bound to the promoter region of the NnF3’H gene. Our study provides in planta functional validation of genes governing flavonoid biosynthesis and regulation specifically within lotus plumule. These findings establish a theoretical framework for further exploration of flavonoid biosynthetic pathways in lotus and offer promising targets for breeding high-flavonoid lotus cultivars.