VuMYB1 and VuMYB4 form an activation–inhibition system that synergistically regulates anthocyanin biosynthesis in bog bilberry (Vaccinium uliginosum) fruit

Color is closely linked to the biosynthesis of anthocyanins, and transcription factors influence anthocyanin accumulation by regulating the expression of structural genes. Anthocyanin biosynthesis is mainly activated by the MBW (MYB-bHLH-WD40) complex, and is co-regulated by multiple inhibitory proteins. Bog bilberry, as a wild blueberry, is rich in anthocyanins, but the molecular mechanisms regulating their biosynthesis remain unclear. Based on transcriptome data, we identified the anthocyanin activator VuMYB1 and the inhibitor VuMYB4. Overexpression of VuMYB1 induces anthocyanin accumulation in bog bilberry fruit and promotes transcription by binding to the promoters of structural genes involved in anthocyanin biosynthesis. Meanwhile, VuMYB1 forms an MBW complex with VubHLH4 and VuTTG1 to regulate anthocyanin production. VuMYB4 interacts with VuMYB1 to form a complex that inhibits anthocyanin synthesis. VuMYB4 directly binds to the promoters of structural genes involved in anthocyanin biosynthesis and suppresses VuMYB1 activity on these promoters. Additionally, VuMYB1 influences the transcription of VuMYB4 by modulating its promoter activity, thereby preventing excessive anthocyanin accumulation. This study suggests that the VuMYB4 and VuMYB1-VubHLH4-VuTTG1 complexes together constitute an "activation-inhibition" system that regulates anthocyanin synthesis in bog bilberry and prevents its excessive accumulation.