OsRbohB-mediated H2O2 signaling underlies rice copper tolerance by regulating copper uptake and detoxification gene expression

Copper (Cu), an essential micronutrient for plant growth, becomes toxic at excessive levels, posing risks to both plant development and human health. Rice ( Oryza sativa L.) as a staple crop for over two-thirds of the global population, requires precise regulation of copper homeostasis to mitigate its toxicity. Hydrogen peroxide (H 2 O 2 ) has been implicated in be involved in the tolerance of rice to copper stress. However, as the major source of reactive oxygen species (ROS), the role and mechanism of respiratory burst oxidase homologs (Rbohs) in the copper stress response of rice remain unclear. In this study, we found that CuSO 4 stress strongly induced OsRbohB expression at both transcriptional and translational levels in rice shoots and roots. Phenotypic and physiological analysis showed that overexpression of OsRbohB significantly enhanced copper stress tolerance, improved photosynthetic capacity, and promoted seedling growth under CuSO 4 stress. Furthermore, OsRbohB was found to positively regulate the increase in H 2 O 2 accumulation induced by CuSO 4 in both shoots and roots. Additionally, OsRbohB suppressed copper uptake by regulating the expression of Cu transporters-related genes ( OsCOPT1, OsHMA5, and OsZIP1 ) and alleviated intracellular copper toxicity by upregulation of metal chelation genes ( OsPCS1 and OsMT-1 ) in roots. Taken together, our findings reveal that OsRbohB modulates Cu homeostasis by coordinating copper uptake and detoxification-related gene expression through H 2 O 2 signaling, thereby enhancing copper tolerance in rice.