Abstract: Soil salinization severely inhibits plant growth, necessitating adaptive changes in root system architecture to enhance stress resilience. Cytokinin signaling plays a key role in regulating root plasticity under salt stress. Type-A response regulators, which lack DNA-binding domains, fine-tune cytokinin signaling through phosphorylation-dependent interactions or transcriptional repression. Although type-A response regulators are associated with stress adaptation, their specific mechanistic roles in salt tolerance remain unclear. Here, we identify PuRR9, a type-A response regulator in poplar, as a negative regulator of cytokinin signaling that enhances salt tolerance through a phosphorylation cascade (PuHK2-PuHP5-PuRR9). Salt stress promotes PuRR9 phosphorylation, strengthening its interaction with the transcriptional repressor PuZFP1. Phosphorylated PuRR9 recruits PuZFP1 to the promoter of PuIPT3, a key cytokinin biosynthesis gene, thereby suppressing its expression. This repression reduces cytokinin biosynthesis and promotes root growth under salt stress. We propose that the PuHK2-PuHP5-PuRR9-PuZFP1 module enhances salt tolerance by establishing a phosphorylation-dependent negative feedback loop within the cytokinin pathway. These findings reveal a novel mechanism by which a type-A response regulator modulates root system architecture and extend current understanding of cytokinin signaling.

Key words: cytokinin signaling, phosphorylation, populus ussuriensis, PuRR9, root, salt