Nitrogen modulates carbon-nitrogen metabolism and nutrient stoichiometry to mitigate ozone stress in Quercus aliena

Tropospheric ozone (O₃) is a pervasive phytotoxic air pollutant, and its co-occurrence with increasing nitrogen (N) deposition poses an emerging hazard to forest health and carbon (C)-nitrogen balance. Understanding how trees respond to these interacting stressors is critical for predicting ecosystem resilience under global change. Here, we investigated the physiological, stoichiometric, and molecular mechanisms underlying Quercus aliena responses to elevated O₃ (160 ppb) and N addition (150 kg·ha^-1·yr^-1), alone and in combination, using an integrated approach combining growth traits, photosynthesis, nutrient analysis, and multi-omics profiling. Elevated O₃ severely inhibited chlorophyll synthesis, photosynthetic rate, and root biomass, and depleted key soluble sugars, indicating C assimilation failure. N addition alone enhanced photosynthetic performance, starch and sucrose accumulation, and biomass production. Strikingly, under combined O₃ and N exposure, seedlings exhibited partial recovery of growth and photosynthesis, reduced leaf C:N ratios, and increased N:P ratios, reflecting adaptive nutrient reallocation. Transcriptomic-metabolomic integration revealed that N supplementation under O₃ stress reprogrammed carbohydrate metabolism, up-regulating genes for sugar mobilization and osmoprotectant biosynthesis while down-regulating those for starch storage and glycolysis. Simultaneously, N assimilation shifted toward low-carbon-cost ammonium pathways, with increased glutamine synthetase and glutamate synthase activities and reduced nitrate reductase activity. These coordinated adjustments enhanced soluble C availability, optimized N utilization, and stabilized nutrient stoichiometry, collectively mitigating O₃-induced metabolic disruption. Our findings identify nutrient-driven metabolic plasticity as a key defense against atmospheric pollutant stress, providing mechanistic insight for targeted nutrient management to safeguard forest productivity in increasingly O₃-polluted environments.