The AaMYB61-AaNAC3 Transcriptional Cascade Synergistically Promotes Fruit Softening by Activating Cell Wall...

Compared with traditional fuzzy kiwifruit (Actinidia chinensis), kiwiberry (A. arguta) undergoes markedly rapid softening after harvest, representing an accelerated climacteric ripening process where ethylene plays a central role. To dissect the regulatory network underlying this process, we used the ethylene inhibitor 1-methylcyclopropene (1-MCP) to retard softening and capture potential transcriptional regulators. Through integrated physiological, biochemical, and transcriptomic analyses, we characterised an R2R3-type MYB transcription factor AaMYB61, whose expression is suppressed by 1-MCP. Dual-luciferase reporter and electrophoretic mobility shift assays demonstrated that AaMYB61 directly binds to the promoters of cell wall degradation-related genes (AaPME1 and AaMAN1), thereby promoting pectin and hemicellulose disassembly. Furthermore, AaMYB61 transactivates the NAC transcription factor AaNAC3, with which it physically interacts to synergistically enhance downstream gene expression. Transient overexpression of AaMYB61 or AaNAC3 in kiwiberry accelerates fruit softening, while stable overexpression of AaMYB61 in kiwifruit (A. chinensis) alters leaf cell wall composition. Likewise, heterologous overexpression of AaMYB61 in tomato accelerates fruit softening and upregulates the expression of ripening-associated genes. These findings reveal that the AaMYB61-AaNAC3 module, which is suppressed by 1-MCP, acts as a transcriptional hub regulating cell wall disassembly and fruit softening in kiwiberry, providing molecular insights for fruit quality improvement and preservation strategies.