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Ethylene regulates anthocyanin synthesis in ripening apple fruit via the antagonistic activities of the R2R3-MYB repressors and activators. However, the molecular mechanism underlying this process remains unknown. In this study, ethylene significantly induced the expression of the R2R3-MYB gene MdMYB17 in apple fruit. Moreover, MdMYB17 was revealed to be an important repressor of anthocyanin synthesis. Specifically, MdMYB17 binds directly to the promoters of the ethylene-induced genes MdMYB1 and MdEIL1, which encode positive regulators of anthocyanin synthesis, and represses their expression. Additionally, MdMYB1 and MdEIL1 bind to the MdMYB17 promoter to activate its expression. Thus, MdMYB17, MdMYB1, and MdEIL1 form a regulatory module that controls the expression of the corresponding genes. MdMYB17 interacts with MdEIL1. The interaction between MdMYB17 and MdEIL1 attenuates the regulatory effects of MdMYB17 on MdMYB1 and MdEIL1 as well as the regulatory effects of MdEIL1 on MdMYB17. Overall, our results reveal the molecular mechanisms by which MdMYB17, MdMYB1, and MdEIL1 finely mediate ethylene-regulated anthocyanin synthesis in apple fruit.

Ethylene precisely regulates anthocyanin synthesis in apple via a module comprising MdEIL1, MdMYB1, and MdMYB17