DcH3.3 and DcNAC1 Regulate the Expression of UGT73A93 Involved in the Changes in Flower Colour and Fungal Resistance in Carnation.

Carnation (Dianthus caryophyllus) contains abundant flavonoid glycosides (FGs), which are important natural functional and colour components. However, there are few reports on the modification of UDP-glycosyltransferases (UGTs) in relation to flavonoids in carnation. In this study, we cloned and characterised a flavonoid 3'-O-glucosyltransferase (UGT73A93) in carnation in vitro. Overexpression of UGT73A93 in carnation and tobacco increased flavonoid glycoside accumulation, particularly kaempferol glycosides, while decreasing anthocyanin content and lightening flower colour. UGT73A93 also enhanced fungal resistance, antioxidant capacity, anti-amylase and anti-pancreatic lipase activities. Yeast one-hybrid and dual-luciferase assays revealed that the UGT73A93 promoter interacted with DcH3.3 and DcNAC1, key regulatory proteins involved in flavonoid biosynthesis. We predicted the interaction between DcLON2 and DcNAC1 using AlphaFold 3 and confirmed this hypothesis through yeast two-hybrid assay and bimolecular fluorescence complementation assays. These findings suggest an epigenetic-transcriptional cascade (DcH3.3-DcNAC1-UGT73A93) wherein DcH3.3 opens chromatin for DcNAC1-mediated UGT73A93 activation, while DcLON2 potentially degrades DcNAC1 to form a feedback loop. These results provide new insights into flavonoid 3'-O-glucosyltransferase and may contribute to future strategies aimed at improving the benefits of flavonoid biosynthesis for both plants and humans. It also demonstrates that AI can be applied in the field of plant biosynthesis, accelerating the process of plant breeding.