The composition and function of root exudates in rhizosphere iron (Fe) mobilization are significantly influenced by environmental pH conditions. While the role of organic acids in Fe solubilization is well-recognized, the molecular mechanisms underlying this pH-dependent process remain poorly understood. Here, we demonstrate that under weakly acidic conditions, proton excretion alone is insufficient to solubilize sparingly soluble Fe. Instead, a pH-dependent ligand specificity emerges as a critical factor in Fe mobilization. Notably, within the pH range of 5.0–6.0, citric acid exuded by roots exhibits superior Fe solubilization efficacy compared to oxalic acid and malic acid. We identified SlFRDL1, a gene induced by Fe deficiency, as a key player in this process. SlFRDL1 encodes a plasma membrane-localized protein with citrate permeability, as confirmed by its functional expression in Xenopus oocytes. Knockout mutants of SlFRDL1 displayed exacerbated Fe deficiency symptoms, which were associated with a significant reduction in citrate secretion from roots. Furthermore, we discovered that SlSTOP1, a transcription factor, binds to the promoter region of SlFRDL1 and activates its expression. Slstop1 mutants exhibited leaf chlorosis symptoms similar to those observed in Slfrdl1 mutants, highlighting the functional interplay between these two genes. Interestingly, while Fe deficiency triggers the FER-mediated Fe uptake system under both acidic and alkaline conditions, the SlSTOP1–SlFRDL1 module is specifically activated only in acidic environments. This pH-specific regulation underscores the importance of the SlSTOP1–SlFRDL1 pathway in root-mediated Fe solubilization under acidic conditions.

citrate exudation  /  iron (Fe) mobilization  /  pH  /  SlFRDL1  /  SlSTOP1