An R3-MYB gene of Phalaenopsis, MYBx1, represses anthocyanin accumulation

Anthocyanins are secondary metabolites that are involved in providing flower petal pigmentation. In plants, anthocyanin synthesis is regulated positively by a conserved MYB-bHLH-WD40 (MBW) complex and negatively by repressors. We isolated a repressor of anthocyanin synthesis, MYBx1, from a moth orchid Phalaenopsis cv. Big Chili. MYBx1 encodes a typical R3-MYB protein and exhibited diverse expression patterns in four Phalaenopsis cultivars, Big Chili, Fuller’s Sunset, Sogo Yukidian “V3”, and Sogo Lit-Sunny. A yeast two-hybrid assay analysis indicated that MYBx1 can also participate in the MBW complex by interacting with the transcription factors ANTHOCYANIN (AN) 1 and AN11. Over-expressing MYBx1 in the petunia hybrid M1 × R27 resulted in reduced accumulations of flower petal pigments through the repression of anthocyanin activities. A quantitative real-time PCR analysis of the expression patterns of anthocyanin synthesis-related structural genes in the transgenic petunia flower petals showed that the expression levels of chalcone synthase, flavanone 3-hydroxylase, and dihydroflavonol reductase genes were considerably down-regulated. However, no significant changes in the expression levels of the MBW complex members, AN1, AN2, and AN11, were observed. This study improves our understanding of anthocyanin synthesis-related regulatory networks in plants and also provides an important foundation for the further genetic engineering of ornamental plant flower colors.