BnaC03.MYB56 and BnaA09.MYB61 antagonistically regulate flavonoid metabolism in Brassica napus

Flavonoid biosynthesis is negatively correlated with seed oil content in B. napus. However, the molecular mechanisms of flavonoid metabolism mediated by MYB repressor in B. napus are still not well understood. In this study, we identify an R2R3-MYB transcription factor, BnaC03.MYB56, which regulates flavonoid metabolism and oil content in B. napus. Overexpression of BnaC03.MYB56 reduced flavonoid metabolite levels and significantly increased seed oil content by 11.0%-11.4% compared to the wild-type, whereas the mutant lines exhibited the opposite phenotype. BnaC03.MYB56 directly binds to the "TAACGA" motif in the promoters of BnaC08.TT6 which catalyzes the synthesis of dihydrokaempferol, and represses its expression. In addition, BnaA09.MYB61 interacts with BnaC03.MYB56 and antagonizes its transcriptional repression of the BnaC08.TT6. BnaA09.MYB61 positively regulates flavonoid metabolism and oil content through activating the expression of BnaC08.TT6. Collectively, this study develops an antagonistic BnaC03.MYB56-BnaA09.MYB61 module that fine-tunes flavonoid metabolism to modulate seed oil content, providing a new target for improving oil content in B. napus breeding.