Multilayered regulation of Fe excess stress and the role of ascorbate in rice

Ascorbate, a vital water-soluble antioxidant, plays a central role in scavenging reactive oxygen species (ROS) in plants under stress conditions, including iron (Fe) toxicity. Iron toxicity, primarily caused by excess ferrous ions (Fe²⁺) in flooded paddy fields, significantly limits rice (Oryza sativa L.) productivity, a staple crop for nearly two-thirds of the global population. This study explores the role of ascorbic acid in regulating iron stress responses in two contrasting rice genotypes-IR64 (sensitive) and Lachit (tolerant). Integrated physiological, biochemical, histochemical, metabolomic, and molecular analyses revealed that Lachit showed reduced Fe accumulation, lower lipid peroxidation, and enhanced ROS scavenging under Fe excess. Notably, the upregulation of key ascorbate biosynthesis genes (VTC1-3, VTC4, VTC2-like) in Lachit suggests a positive correlation between ascorbic acid production and Fe stress tolerance. Histochemical staining demonstrated higher hydrogen peroxide (H₂O₂) scavenging in Lachit, confirming better ROS detoxification. Metabolomic profiling showed elevated ascorbate levels and a suite of antioxidant metabolites contributing to improved stress resilience. Additionally, higher accumulation of secondary metabolites such as polyphenols and flavonoids across both genotypes suggests a synergistic role with ascorbate in mitigating Fe-induced oxidative stress. Collectively, these findings highlight ascorbate as a key regulator in iron stress mitigation and underscore its interaction with secondary metabolites in strengthening antioxidative defense. This study provides novel insights into ascorbate-centered mechanisms and identifies promising molecular targets for improving iron excess tolerance and productivity in rice through genetic engineering or genome editing approaches.