PeCPK21 interacts with HMAPs to tolerate elevated cadmium stress in Populus × canescens

Improving Cd 2+ tolerance in poplars is crucial for reclaiming Cd 2+ -contaminated soils with fast-growing trees. Overexpression of PeCPK21 , a Ca 2+ -dependent protein kinase gene from Populus euphratica , mitigates the toxicity of elevated CdCl 2 stress (500 µM, 30 d) in Populus × canescens . HaloTag pull-down and mass spectrometry assays showed that PeCPK21 interacted with various heavy metal stress-associated proteins (HMAPs), including heavy metal transport-related proteins, photosynthesis-linked proteins, membrane intrinsic proteins, and antioxidant enzymes. The corresponding expression profiles of HMAPs following cadmium exposure revealed that the PeCPK21-interacting proteins served to limit the build-up of Cd 2+ , boost antioxidant defenses, and improve photosynthesis and water management in Cd 2+ -stressed poplars. PcXTH6 and PcSOD[Fe] were transferred to P. × canescens and Arabidopsis to confirm the contribution of PeCPK21-interacting proteins to Cd 2+ tolerance. Overexpression of PcXTH6 increased the ability to limit Cd 2+ in transgenic P. × canescens and Arabidopsis under elevated cadmium stress. This was due to the PcXTH6-enhanced xyloglucan degradation activity, which reduces Cd 2+ binding sites in the cell wall, ultimately leading to decreased Cd 2+ uptake and accumulation under high cadmium conditions. Furthermore, PcSOD[Fe] overexpression increased ROS-scavenging capacity in transgenic P. × canescens and Arabidopsis under elevated cadmium stress. It can be concluded that PeCPK21 interacts with PcXTH6 and PcSOD[Fe] to limit Cd 2+ and ROS concentrations under prolonged and severe cadmium exposure, thereby attenuating the detrimental effects of Cd 2+ on plant growth, photosynthesis, water retention, and antioxidant defenses. The insights on interactions between PeCPK21-HMAPs could be used to genetically engineer woody species for Cd 2+ tolerance. • PeCPK21 interacts with HMAPs to mitigate toxicity of high Cd in Populus × canescens . • PeCPK21 interacts with PcXTH6 to limit Cd accumulation in poplar and Arabidopsis. • PeCPK21 interacts with PcSOD[Fe] to enhance ROS scavenging in poplar and Arabidopsis. • PeCPK21 interacts with photosynthesis-linked proteins to retain carbon assimilation. • PeCPK21 interacts with membrane intrinsic proteins to improve water status in poplar.