A novel R2R3‐MYB transcription factor FaMYB5 positively regulates anthocyanin and proanthocyanidin biosynthesis in cultivated strawberries (Fragaria × ananassa)

Abstract

Flavonoids have a major contribution to the fruit quality in cultivated strawberries and are regulated by MYB, bHLH and WD40 transcriptional factors. We reported here the identification of the FaMYB5, an R2R3-MYB transcription factor, which positively regulated the accumulation of anthocyanins and proanthocyanidins through the trans-activation of the F3'H and LAR. The strawberry FaEGL3 and FaLWD1/FaLWD1-like interact with the R2R3-FaMYB5 to form an MYB-bHLH-WD40 complex (MBW), enhancing the regulatory efficiency. The R2R3-FaMYB5 was constitutively expressed in various tissues and in fruits of different developmental stages, which was strikingly contrasting to the fruit-specific expression patterns of FaMYB10. Meanwhile, R2R3-FaMYB5 failed to promote a stable accumulation of anthocyanin glycosides in the mature fruits of the myb10 mutant, mainly due to the suppressed expression of TT19. The R2R3-FaMYB5 was regulated by an antisense long noncoding RNA lncRNA-myb5. Additionally, the R2R3-FaMYB5 protein could interact with FaBT2 and was degraded through the ubiquitin/26 S proteasome pathway. Transcriptome and metabolome data showed that R2R3-FaMYB5 enhanced the gene expression and the metabolite accumulation involved in the flavonoid, phenylpropanoid and lignin biosynthesis pathways. Collectively, we conclude that the FaMYB5 is an R2R3-MYB activator involved in the composition of MBW, which positively regulates the biosynthesis of anthocyanin and proanthocyanidin. These findings provided new insights into the molecular mechanisms that regulate flavonoids in strawberry fruits.

Regulation of flavonoids in strawberry fruits by FaMYB5/FaMYB10 dominated MYB-bHLH-WD40 ternary complexes

“…As previously reported, R2R3-FaMYB5 had an intact R2 domain compared with the former investigated R3-FaMYB5, another start codon can be found at 83 aa before R3-FaMYB5, which could encode an MYB5 protein with a complete R2R3 domain ( Figures 1A, B ; Schaart et al., 2013 ; Jiang et al., 2023 ). Sequence alignment results indicated that FaMYB1, R2R3-FaMYB5, FaMYB9, FaMYB10, and FaMYB11 belong to R2R3-type MYB TFs, in which FaMYB1 had an EAR repression motif (pdLNLD/EL) of flavonoids on the C2 domain ( Figure 1A ; Aharoni et al., 2001 ).…”

“…In our previous report ( Jiang et al., 2023 ), it was preliminarily elucidated the interaction between FaMYB5, FaEGL3, FaLWD1 and FaLWD1-like proteins. Here, we expanded the test gene list to get the full landscape of the MBW complex in strawberry.…”

“…Surprisingly, no fluorescent signals were detected between FabHLH3/FaGL3/FaTTG1 and FaMYB5/10. The signals for FaMYB5/10 and FaMYC1 proteins were faint, while other positive interactions could still be observed by BiFC (Figure 3B; Jiang et al, 2023). Therefore, we mainly explored the consistent interactions selected by Y2H and BiFC in the following experiments, especially the FaMYB5/10, FaEGL3, and FaLWD1/ FaLWD1-like TFs that had the potential ability to form the MBW complex (Figure 3C).…”

“…In previous study, we used comparative transcriptomics and metabolomics to evaluate the effects of FaMYB5 on strawberry flavonoid and flavonoid-related pathway, further explored the regulation of R2R3-FaMYB5 via lncRNAs and FaBT2 at the transcription and protein levels ( Jiang et al., 2023 ). Herein, in order to get the full landscape of the MBW complex in strawberry, we took advantage of molecular methods to screen the interacting members that may form the MBW complex and regulate the flavonoids in strawberry fruits.…”

SlMYB7, an AtMYB4-Like R2R3-MYB Transcription Factor, Inhibits Anthocyanin Accumulation in Solanum lycopersicum Fruits

“…11,12 The MBW ternary complex, which is essential in regulating the expression of genes related to anthocyanin synthesis, comprises a MYB, a bHLH, and a WD40 TF. 13,14 For example, R2R3-MYBs in the MBW recognize and bind to target gene promoters, directly affecting anthocyanin accumulation by determining the expression levels and tissue specificity of downstream structural genes. 15,16 In addition, two distinct groups of MYB TFs, the CPC-MYB (R3-MYB) 17−19 and R2R3-MYB repressors, 20,21 also play significant roles in the regulatory network controlling anthocyanin biosynthesis.…”