Abscisic acid-responsive transcription factors PavDof2/6/15 mediate fruit softening in sweet cherry

Softening is a key step during fruit ripening that is modulated by the interplay between multiple phytohormones. The antagonistic action of abscisic acid (ABA) and auxin determines the rate of fruit ripening and softening. However, the transcription factors that integrate ABA and auxin signals to regulate fruit softening remain to be determined. In this study, we identified several DNA-binding with One Finger (Dof) transcription factors essential for ABA-promoted fruit softening, based on transcriptome analysis of two sweet cherry (Prunus avium L.) varieties with different fruit firmness. We show that PavDof6 directly binds to the promoters of genes encoding cell wall-modifying enzymes to activate their transcription, while PavDof2/15 directly repress their transcription. Transient overexpression of PavDof6 and PavDof2/15 in sweet cherry fruits resulted in precocious and delayed softening, respectively. In addition, we show that the auxin response factor PavARF8, the expression of whose encoding gene is repressed by ABA, activates PavDof2/15 transcription. Furthermore, PavDof2/6/15 and PavARF8 directly bind to the 9-cis-epoxycarotenoid dioxygenase 1 (PavNCED1) promoter and regulate its expression, forming a feedback mechanism for ABA-mediated fruit softening. These findings unveil the physiological framework of fruit softening and establish a direct functional link between the ABA-PavARF8-PavDofs module and cell-wall-modifying genes in mediating fruit softening.