Glutathione-associated genes SpGSH1 and SpPCS1 differentially modulate cadmium accumulation and tolerance in duckweed under micronutrient deficient environments.

Decontaminating cadmium (Cd) pollution, a severe threat to aquatic ecosystems, requires efficient phytoremediation solutions. Duckweed (Spirodela polyrhiza) exhibits Cd bioremediation potential, but the roles of glutathione (GSH)-associated genes under Cd and iron/manganese (Fe/Mn) deficiency stress remain unclear, limiting its practical applications. To address this, we generate transgenic lines overexpressing SpGSH1 (G line), SpPCS1 (P line), and both SpGSH1/SpPCS1 (GP line) and investigate their roles in Cd accumulation and tolerance under these three stress conditions. We report elevated GSH levels and total antioxidant capacity in all transgenic lines, but only P and GP lines significantly enhance Cd accumulation under 20 μM Cd stress. Under combined Fe deficiency and 20 μM Cd stress, SpGSH1 overexpression impairs growth, whereas SpPCS1 overexpression causes severe photosynthetic damage. Only the GP line restores Cd accumulation capacity and effectively mitigates oxidative stress, indicating a synergistic detoxification mechanism. Under combined Mn deficiency and 20 μM Cd stress, nutrient-specific divergence emerges with the G line, with enhanced growth, improved ROS scavenging, and protected photosynthesis, whereas the P line accumulates O2- levels. Simultaneously, the GP line maintains robust antioxidant protection while boosting Cd/Zn uptake. Importantly, SpGSH1/SpPCS1 co-overexpression synergistically improves Cd accumulation, antioxidant capacity, and photosynthesis across stress conditions, overcoming limitations of single-gene overexpression. We pioneer a novel SpGSH1/SpPCS1 co-overexpression strategy for robust Cd remediation in nutrient-depleted waters that represents a promising approach to enhance phytoremediation efficacy in challenging real-world environments.