MdGSTF6, activated by MdMYB1, plays an essential role in anthocyanin accumulation in apple.

Anthocyanins are biosynthesized on the cytosolic surface of the endoplasmic reticulum and then transported into the vacuole for storage. Glutathione S-transferases (GSTs) are considered to be responsible for the transport of anthocyanins into the vacuole. However, the regulatory mechanisms of GSTs in plants are still unclear. Here, we performed a genome-wide analysis and identified 69 GST genes in apple. The expression of MdGSTF6 was positively correlated with the anthocyanin content (r = 0.949) during 'Yanfu 8' fruit development. The overexpression of MdGSTF6 in the Arabidopsis thaliana tt19 mutant resulted in seedlings of 35S::MdGSTF6-GFP/tt19 that could accumulate anthocyanin and rescue its phenotype, suggesting that MdGSTF6 was an anthocyanin transporter. The silencing of MdGSTF6 affected anthocyanin accumulation in apple fruit. Moreover, the knockdown of MdGSTF6 by RNA interference in cultured 'Gala' seedlings inhibited anthocyanin accumulation. The interaction experiments showed that MdMYB1 could bind directly to the MdGSTF6 promoter to transcriptionally activate its expression. Collectively, our results demonstrate that MdGSTF6 encodes an important GST transporter of anthocyanins in apple fruit and provide evidence for the associated regulatory mechanisms. Therefore, MdMYB1 can not only regulate anthocyanin synthesis, but also control the transport of anthocyanin in apples. This information may be useful for further clarifying the regulation of anthocyanin transport in apple.

MdCOL4 Interaction Mediates Crosstalk Between UV-B and High Temperature to Control Fruit Coloration in Apple.

In many plants, anthocyanin biosynthesis is affected by environmental conditions. Ultraviolet-B (UV-B) radiation promotes anthocyanin accumulation and fruit coloration in apple skin, whereas high temperature suppresses these processes. In this study, we characterized a B-box transcription factor, MdCOL4, from 'Fuji' apple, and identified its role in anthocyanin biosynthesis by overexpressing its encoding gene in apple red callus. The expression of MdCOL4 was reduced by UV-B, but promoted by high temperature. We explored the regulatory relationship between heat shock transcription factors (HSFs) and MdCOL4, and found that MdHSF3b and MdHSF4a directly bound to the heat shock element cis-element of the MdCOL4 promoter. MdCOL4 interacted with MdHY5 to synergistically inhibit the expression of MdMYB1, and MdCOL4 directly bound to the promoters of MdANS and MdUFGT, which encode genes in the anthocyanin biosynthetic pathway, to suppress their expression. Our findings shed light on the molecular mechanism by which MdCOL4 suppresses anthocyanin accumulation in apple skin under UV-B and high temperature.