PuMYB40 and PuWRKY75 synergistically enhance phosphate uptake and organic phosphorus hydrolysis under phosphate deficiency in poplar

Inorganic phosphate deficiency severely limits tree growth and productivity. The challenge of phosphorus acquisition extends beyond the scarcity of inorganic phosphate, as the utilization of organic phosphorus represents another critical yet poorly understood adaptation in woody plants. Here, we identify a synergistic regulatory module involving the R2R3-MYB transcription factor PuMYB40 and the WRKY factor PuWRKY75 that enhances organic phosphorus scavenging and inorganic phosphate uptake in Populus ussuriensis under inorganic phosphate-deficient conditions. Overexpression of PuMYB40 or PuWRKY75 enhanced inorganic phosphate acquisition and plant growth when phytate was supplied as the sole phosphorus source or under low-inorganic phosphate conditions, whereas suppression had the opposite effects. RNA sequencing revealed that both factors activate numerous inorganic phosphate starvation-responsive genes, with over 60% of PuMYB40-regulated genes overlapping with those controlled by PuWRKY75. Molecular analyses demonstrated that PuMYB40 and PuWRKY75 directly activate the expression of PuPAP17 (encoding a rhizosphere-secreted acid phosphatase), PuPHT1;4 (a high-affinity inorganic phosphate transporter), PuPHT1;9 (a root-to-shoot inorganic phosphate translocator), and PuGDPD1 (which facilitates phospholipid remodeling) by binding to distinct cis-elements in their promoters. Overexpression of PuPAP17 or PuPHT1;4 increased organic phosphorus hydrolysis when phytate was supplied as the sole phosphorus source and phosphate uptake under low-inorganic phosphate conditions, respectively. Transient dual-luciferase and microscale thermophoresis assays revealed that PuWRKY75 and PuMYB40 interact, and this interaction amplifies the transcriptional activation of PuPAP17 and PuPHT1;4 by PuMYB40 but not by PuWRKY75. Our findings establish a coordinated "decompose-import" regulatory axis in poplar, providing mechanistic insights and genetic tools for improving phosphorus-use efficiency in forest trees.