The deubiquitinase UBP15 and the E3 ubiquitin ligase PUB17 coordinate GLK1 stability in apple anthocyanin deposition

Ubiquitination functions in brassinosteroid (BR) signal transduction and BR regulates anthocyanin biosynthesis; however, the function of deubiquitination in these processes remains unclear. Here, we identified the transcription factor GOLDEN2-LIKE 1 (MdGLK1) as a central regulatory hub for BR-mediated anthocyanin accumulation in apple (Malus × domestica). MdGLK1 relieved the transcriptional repression of the core BR signaling regulator BRASSINAZOLE-RESISTANT 1 (MdBZR1) on MYELOBLASTOSIS 1 (MdMYB1), a positive regulator of anthocyanin biosynthesis, and interacted with MdMYB1 to enhance its transcriptional activation of anthocyanin biosynthetic genes, thereby promoting anthocyanin production. MdGLK1 protein stability was coordinately regulated by the plant U-box type E3 ubiquitin ligase 17 (MdPUB17) and the ubiquitin-specific protease 15 (MdUBP15) in response to BR signaling. BR-activated MdPUB17 promoted MdGLK1 degradation, thereby reducing anthocyanin accumulation promoted by MdGLK1. Conversely, the deubiquitinase MdUBP15 maintained MdGLK1 protein homeostasis and sustained MdGLK1-promoted anthocyanin deposition by counteracting the MdPUB17-mediated ubiquitination and degradation of MdGLK1 in the absence of BRs. In summary, the BR-responsive factor MdGLK1 regulates anthocyanin biosynthesis by modulating the transcriptional activities of MdBZR1 and MdMYB1 and its protein turnover is dynamically controlled by a ubiquitination-deubiquitination switch formed by MdPUB17 and MdUBP15. These findings thus provide insights into the post-translational regulatory network of BR signaling and pigment accumulation.