PrNAC72 Activates PrDXS2 to Regulate Monoterpene Biosynthesis in Paeonia rockii

ABSTRACT Paeonia rockii is a woody ornamental plant of high horticultural value that is widely recognized for its distinctive terpene-rich floral aroma. However, the molecular regulation of terpene production in this species remains largely unclear. Here, dynamic volatile analysis throughout five flower development stages confirmed that monoterpenes are the dominant fragrance components. A combined transcriptomic approach using the Illumina RNA-seq and PacBio Iso-Seq platforms identified 7 254 differentially expressed genes (DEGs), 147 of which were associated with terpenoid metabolic pathways. Notably, 16 DEGs were annotated as isoforms of 1-deoxy-D-xylulose 5-phosphate synthase (DXS), a rate-limiting enzyme in the methylerythritol phosphate (MEP) pathway. Among them, PrDXS1 and PrDXS2 exhibited petal-specific expression trends closely aligned with volatile monoterpene release, and both were localized to plastids. Functional assays, including transient overexpression and virus-induced gene silencing (VIGS), revealed that these genes positively influenced monoterpene accumulation. In addition, the NAC transcription factor PrNAC72 directly regulated PrDXS2 expression, as validated by yeast one-hybrid, dual-luciferase, and GUS reporter experiments. The overexpression of PrNAC72 in heterologous systems confirmed the role of PrNAC72 in promoting monoterpene biosynthesis. These results reveal a novel regulatory pathway involving PrNAC72 and PrDXS2 in floral scent biosynthesis and offer promising molecular targets for enhancing aroma traits in tree peony breeding.