Populus euphratica PeRAX2 interacts with the PeANN1 promoter to regulate gene expression and cadmium tolerance

Poplar species hold considerable potential for large-scale phytoremediation efforts and there is a growing interest in enhancing the Cd²⁺ tolerance of poplars via genetic engineering. PeRAX2, the R2R3-MYB transcription factor from Populus euphratica, facilitates AtANN1 expression and Cd²⁺ uptake in Arabidopsis, but it remains uncertain whether PeRAX2 regulates Cd²⁺ accumulation and tolerance in poplars. When exposed to cadmium stress, P. euphratica increased the transcripts of PeRAX2, while decreasing the expression of PeANN1, which promotes the entry of Cd²⁺ in Arabidopsis. PeRAX2 was identified to interact with the PeANN1 promoter through DNA pull-down, EMSA, Y1H, VIGS, LUC reporter assay, and transient overexpression assay. Intriguingly, PeRAX2 bound to the PeANN1 promoter and repressed the gene expression. Moreover, PeRAX2 was shown to limit Cd²⁺ buildup by inhibiting PeANN1 expression in P. euphratica leaves that transiently overexpressed PeRAX2 when exposed to cadmium. Conversely, PeANN1 overexpression in P. euphratica and P. × canescens resulted in increased Cd²⁺ concentrations, which led to reduced growth, impaired photosynthesis, and weakened antioxidant defenses to detoxify the reactive oxygen species. Hence, the upregulated PeRAX2 in cadmium-challenged P. euphratica leads to the downregulation of PeANN1; this aids the poplars exposed Cd²⁺ to curtail the uptake and build-up of Cd²⁺, thereby fostering the maintenance of plant growth, photosynthesis, and ROS homeostasis in Cd²⁺-stressed environments. These findings regarding distinctive function of PeRAX2 in regulating ANN1 expression in P. euphratica and Arabidopsis can be utilized to genetically modify plant species for targeted Cd²⁺ uptake and Cd²⁺ tolerance.