The <scp>PopGRF3</scp> ‐ <scp>PopKNAT3</scp> module orchestrates salt tolerance through transcriptional control of stress‐responsive genes in poplar

Plants are often exposed to various stressors during their growth and development. Sustaining intracellular ion equilibrium, especially the balance between Na⁺ and K⁺, plays a vital role in plant adaptation to salt stress. In this study, we identified growth-regulating factor 3 (PopGRF3), a gene encoding a transcription factor (TF) that acts as a negative regulator of salt stress responses. PopGRF3-overexpressing (OE) and PopGRF3-RNA interference (RNAi) lines exhibited increased sensitivity and enhanced tolerance to salt stress, respectively. Notably, when exposed to salt stress, PopGRF3 interacts with the promoter region of the sodium/hydrogen exchanger 2 (PopNHX2), leading to the suppression of its transcriptional activity. It is demonstrated that knotted-like homeobox 3 (PopKNAT3), a gene encoding a TF, physically interacts with PopGRF3 and mitigates the latter's inhibitory effect on PopNHX2. Although PopKNAT3 does not directly regulate PopNHX2 expression, the PopKNAT3-PopGRF3 protein complex prevents PopGRF3 from binding to the PopNHX2 promoter. Consequently, transgenic plants overexpressing PopKNAT3 exhibited improved salt tolerance, while those with RNAi-mediated silencing of PopKNAT3 showed increased salt sensitivity. The dual role of the PopGRF3-PopKNAT3 module in regulating salt stress reveals a novel mechanism in Populus, thereby offering new insights into the complex networks that control these stress responses.