Functional characterization of FchDOF1, a transcription factor involved in softening of Fragaria chiloensis fruit

Fruit softening is key for strawberry quality and shelf-life. Cell wall structure is closely associated with texture, and its assembly/disassembly takes place during fruit development and ripening. Softening requires the coordinated spatiotemporal action of sets of cell wall modifying enzymes and therefore a transcriptional regulation is expected. Members of DOF (DNA-binding with One Finger) transcription factor (TF) family have been extensively studied in plant development and stress responses; however, their direct involvement in fruit softening remains poorly understood, particularly in non-climacteric fruits. In this study, the functional characterization of FchDOF1 from Fragaria chiloensis was accomplished and its role during fruit ripening was investigated. FchDOF1 localizes in the nucleus and displays a fruit-specific expression pattern, with a marked induction during ripening. Transient modulation of FchDOF1 expression in fruits reveals that overexpression significantly increases transcript levels of key cell wall-related genes, including FchPL, FchPG, FchXTH1 and FchEXP5, whereas RNAi-mediated silencing leads to reduction of FchPL and FchPG transcripts. Dual-luciferase assay further demonstrates that FchDOF1 specifically transactivates the FchPL promoter and EMSA assays confirmed its specific binding to DOF-recognition motifs within the FchPL promoter, supporting a direct regulatory interaction. In silico promoter analysis identified differential distributions of DOF-responsive elements among cell wall genes, suggesting promoter architecture-dependent regulatory specificity. Together, these results provide functional evidence of FchDOF1 acting as positive regulator of a subset of genes involved in cell wall disassembly during fruit ripening. This study expands current knowledge on functional diversification of DOFs and highlights their contribution to fruit softening.