An R2R3-MYB Transcription Factor RoMYB10 Regulates Anthocyanin Biosynthesis in Black Raspberry

Abstract

At present, there are many excellent cultivars of raspberry worldwide with different fruit colors, such as yellow, red, purple and black. Anthocyanin accumulation is responsible for flower and fruit coloration in plants. Anthocyanins belong to flavonoids which are natural pigments widely distributed in plants. The MYB transcription factor (TF) plays a vital regulatory role in the metabolic pathway of anthocyanins. Here, RoMYB10, an R2R3-MYB in black raspberry (Rubus occidentalis L.), was isolated. The basic physical and chemical properties, conserved motif distribution, evolutionary relationship and subcellular localization were analyzed for the RoMYB10 encoded amino acid sequence. The contents of anthocyanin were determined in four different colored raspberry fruits by high-performance liquid chromatography–tandem mass spectrometry (HPLC-MS). The expression levels of MYB10 genes in various tissues and fruits at different developmental stages of the four different colored raspberries were detected by real-time fluorescence quantitative PCR (qRT-PCR). The results showed that the contents of anthocyanin and the expression levels of the MYB10 gene were the highest in black raspberry. Moreover, the expression of MYB10 in the fruit was significantly higher than in other tissues, especially in ripe and mottled fruits. When ectopic overexpression of RoMYB10 occurred in tobacco, the expression levels of the RoMYB10 gene and total anthocyanin contents in transgenic tobacco plants were significantly higher than those in wild-type (WT) plants. In addition, overexpression of RoMYB10 up-regulated the key genes in the anthocyanin biosynthesis, such as NtCHS, NtCHI, NtF3H, NtANS and NtUFGT. Our study suggests that RoMYB10 is involved in the regulating of anthocyanin biosynthesis in raspberry and has potential as a molecular tool for manipulating biosynthesis of anthocyanins in fruits using metabolic engineering.