ABSTRACT
R3-MYBs negatively regulate anthocyanin pigmentation in plants. However, how R3-MYB repressors finely modulate anthocyanin biosynthesis in cooperation with R2R3-MYB activators remains unclear in monocots. We previously identified two anthocyanin-related R2R3-MYB activators (MaMybA and MaAN2) in grape hyacinth (Muscari spp.). Here, we isolated a R3-MYB repressor, MaMYBx, and characterized its role in anthocyanin biosynthesis using genetic and biochemical markers. The temporal expression pattern of MaMYBx was similar to that of MaMybA and MaAN2, and it was correlated with anthocyanin accumulation during flower development. MaMYBx could be activated either by MaMybA alone or by MaMybA/MaAN2 and cofactor MabHLH1, and it suppressed its own activation and that of MaMybA promoters mediated by MaMybA/MaAN2 and MabHLH1. Like MaMybA, MaMYBx interacted with MabHLH1. MaDFR and MaANS transcription and anthocyanin accumulation mediated by MaMybA/MaAN2 and MabHLH1 were inhibited by MaMYBx. Overexpression of MaMYBx in tobacco greatly reduced flower pigmentation and repressed the expression of late structural and regulatory anthocyanin pathway genes. Thus, MaMYBx finely regulates anthocyanin biosynthesis by binding to MabHLH1 and disrupting the R2R3 MYB-bHLH complex in grape hyacinth. The regulatory network of transcriptional activators and repressors modulating anthocyanin biosynthesis is conserved within monocots. MaMYBx seems a potentially valuable target for flower color modification in ornamental plants.
Subject(s)
Anthocyanins/biosynthesis , Asparagaceae/genetics , Gene Expression Regulation, Plant , Pigments, Biological/metabolism , Plant Proteins/genetics , Amino Acid Sequence , Anthocyanins/genetics , Asparagaceae/metabolism , Phylogeny , Pigments, Biological/genetics , Plant Proteins/chemistry , Plant Proteins/metabolism , Plants, Genetically Modified/genetics , Plants, Genetically Modified/metabolism , Sequence Alignment , Nicotiana/genetics , Nicotiana/metabolismABSTRACT
Flavonols are important copigments that affect flower petal coloration. Flavonol synthase (FLS) catalyzes the conversion of dihydroflavonols to flavonols. In this study, we identified a FLS gene, MaFLS, expressed in petals of the ornamental monocot Muscari aucheri (grape hyacinth) and analyzed its spatial and temporal expression patterns. qRT-PCR analysis showed that MaFLS was predominantly expressed in the early stages of flower development. We next analyzed the in planta functions of MaFLS. Heterologous expression of MaFLS in Nicotiana tabacum (tobacco) resulted in a reduction in pigmentation in the petals, substantially inhibiting the expression of endogenous tobacco genes involved in anthocyanin biosynthesis (i.e., NtDFR, NtANS, and NtAN2) and upregulating the expression of NtFLS. The total anthocyanin content in the petals of the transformed tobacco plants was dramatically reduced, whereas the total flavonol content was increased. Our study suggests that MaFLS plays a key role in flavonol biosynthesis and flower coloration in grape hyacinth. Moreover, MaFLS may represent a new potential gene for molecular breeding of flower color modification and provide a basis for analyzing the effects of copigmentation on flower coloration in grape hyacinth.
