Conclusion
Our findings demonstrate that a MaXB3-MaNAC029 module regulates ethylene biosynthesis and a series of cellular metabolisms related to fruit quality formation during banana ripening. These results expand the understanding of the transcriptional and post-translational mechanisms of fruit ripening and quality formation.
Methods
We used RNA-seq to compare unripe and ripe banana fruits and identified a ripening-induced NAC transcription factor, MaNAC029. We further performed DNA affinity purification sequencing to identify the MaNAC029's target genes involved in ethylene biosynthesis and fruit quality formation, and electrophoretic mobility shift assay, chromatin immunoprecipitation with real-time polymerase chain reaction and dual luciferase assays to explore the underlying regulatory mechanisms. Immunoprecipitation combined with mass spectrometry, yeast two-hybrid assay, and bimolecular fluorescence complementation assay were used to screen and verify the proteins interacting with MaNAC029. Finally, the function of MaNAC029 and its interacting protein associated with ethylene biosynthesis and quality formation was verified through transient overexpression experiments in banana fruits.
Results
The study identified a nucleus-localized, ripening-induced NAC transcription factor MaNAC029. It transcriptionally activated genes associated with ethylene biosynthesis and a variety of cellular metabolisms related to fruit quality formation (cell wall degradation, starch degradation, aroma compound synthesis, and chlorophyll catabolism) by directly modulating their promoter activity during ripening. Overexpression of MaNAC029 in banana fruits activated ethylene biosynthesis and accelerated fruit ripening and quality formation. Notably, the E3 ligase MaXB3 interacted with and ubiquitinated MaNAC029 protein, facilitating MaNAC029 proteasomal degradation. Consistent with this finding, MaXB3 overexpression attenuated MaNAC029-enhanced ethylene biosynthesis and quality formation.