The regulatory module MdPUB29-MdbHLH3 connects ethylene biosynthesis with fruit quality in apple

The plant hormone ethylene is critical for climacteric fruit ripening, while glucose and anthocyanins determine the fruit quality of climacteric fruits such as apple. Understanding the exact molecular mechanism for this process is important for elucidating the interconnection of ethylene and fruit quality. Overexpression of apple MdbHLH3 gene, an anthocyanin-related basic helix-loop-helix transcription factor (bHLH TF) gene, promotes ethylene production, and transgenic apple plantlets and trees exhibit ethylene-related root developmental abnormalities, premature leaf senescence, and fruit ripening. Biochemical analyses demonstrate that MdbHLH3 binds to the promoters of three genes that are involved in ethylene biosynthesis, including MdACO1, MdACS1, and MdACS5A, activating their transcriptional expression, thereby promoting ethylene biosynthesis. High glucose-inhibited U-box-type E3 ubiquitin ligase MdPUB29, the ortholog of Arabidopsis AtPUB29 in apple, influences the expression of ethylene biosynthetic genes and ethylene production by direct ubiquitination of the MdbHLH3 protein. Our findings provide new insights into the ubiquitination of MdbHLH3 by glucose-inhibited ubiquitin E3 ligase MdPUB29 in the regulation of ethylene biosynthesis as well as indicate that the regulatory module MdPUB29-MdbHLH3 connects ethylene biosynthesis with fruit quality in apple.