The transcription factors DOF4.6 and XND1 jointly regulate root hydraulics and drought responses in Arabidopsis

Water uptake by roots is essential for plant growth and stress acclimation. We previously showed that the XYLEM NAC DOMAIN 1 (XND1) transcription factor negatively regulates root hydraulic conductivity (Lpr) in Arabidopsis (Arabidopsis thaliana). Here, we show that XND1 physically interacts with the transcription factor DNA-binding with One Finger 4.6 (DOF4.6). Analyses of loss-of-function mutants and overexpression lines revealed that, similar to XND1, DOF4.6 negatively regulates Lpr. DOF4.6 and XND1 jointly modulate downstream gene expression, inhibiting root xylem formation. Notably, DOF4.6 facilitates XND1 binding to the promoter of XYLEM CYSTEINE PROTEASE 1, the product of which controls programmed cell death during xylem development. DOF4.6 also independently binds to the promoters of the aquaporins PIP2;5 and PIP2;6, thereby suppressing their expression and exerting potential direct regulatory effects on membrane water transport. Importantly, the dof4.6 loss-of-function alleles showed significantly enhanced resistance to drought stress. Collectively, our findings demonstrate that DOF4.6 plays a crucial role in root hydraulics and drought stress responses, partially in concert with XND1 and through combined effects on xylem formation and aquaporin functions.