Integrated transcriptomic and metabolomic analyses reveal terpene synthases LlTPS13 and LlTPS14 positively regulate β-ocimene biosynthesis in Lycoris longituba.

The flowers of Lycoris longituba are distinguished by their fragrant petals and dynamic colour variations. Floral volatile compounds play vital roles in plant biology; however, the biosynthetic pathways and regulatory mechanisms underlying fragrance production in L. longituba remain inadequately understood. In this study, metabolomic and transcriptomic analyses were integrated to characterize the variation of 526 volatile organic compounds (VOCs) in L. longituba florets. Metabolite profiling revealed that terpenoids constituted the predominant class of VOCs, while sweet and green aromas were the dominant scent characteristics throughout flower development. Transcriptomic analysis further revealed that differential expression of genes involved in the terpenoid and phenylpropanoid biosynthetic pathways contributed substantially to the regulation of β-ocimene biosynthesis. Weighted gene co-expression network analysis (WGCNA) identified NAC, MADS, HSF, WRKY, bZIP, C2H2, and GARP-G2 transcription factors (TFs) as key regulators of genes closely associated with β-ocimene production. Integrated transcriptomic and correlation analyses further demonstrated that two terpene synthase (TPS) genes were strongly associated with terpenoid biosynthesis across four Lycoris species and throughout floret development. Functional characterization identified the chloroplast-localized terpene synthases LlTPS13 and LlTPS14 as the principal enzymes involved in β-ocimene biosynthesis. Moreover, transient overexpression assays in L. longituba flowers and tobacco plants confirmed that these proteins catalyzed the production of β-ocimene and trans-β-ocimene. These results provide new insights into the variation in floral scent among cultivars and developmental stages and highlight the potential regulatory roles of the identified TPS genes and transcription factors in terpenoid biosynthesis in L. longituba.