Two B-Box Proteins, MaBBX20 and MaBBX51, Coordinate Light-Induced Anthocyanin Biosynthesis in Grape Hyacinth

Floral colour is an important agronomic trait that influences the commercial value of ornamental plants. Anthocyanins are a class of flavonoids and confer diverse colours, and elucidating the molecular mechanisms that regulate their pigmentation could facilitate artificial manipulation of flower colour in ornamental plants. Here, we investigated the regulatory mechanism of light-induced anthocyanin biosynthesis during flower colouration in grape hyacinth (Muscari spp.). We studied the function of two B-box proteins, MaBBX20 and MaBBX51. The qPCR revealed that MaBBX20 and MaBBX51 were associated with light-induced anthocyanin biosynthesis. Both MaBBX20 and MaBBX51 are transcript factors and are specifically localised in the nucleus. Besides, overexpression of MaBBX20 in tobacco slightly increased the anthocyanin content of the petals, but reduced in MaBBX51 overexpression lines. The yeast one-hybrid assays indicated that MaBBX20 and MaBBX51 did not directly bind to the MaMybA or MaDFR promoters, but MaHY5 did. The BiFC assay revealed that MaBBX20 and MaBBX51 physically interact with MaHY5. A dual luciferase assay further confirmed that the MaBBX20-MaHY5 complex can strongly activate the MaMybA and MaDFR transcription in tobacco. Moreover, MaBBX51 hampered MaBBX20-MaHY5 complex formation and repressed MaMybA and MaDFR transcription by physically interacting with MaHY5 and MaBBX20. Overall, the results suggest that MaBBX20 positively regulates light-induced anthocyanin biosynthesis in grape hyacinth, whereas MaBBX51 is a negative regulator.