CITF1 interacts with FIT and regulates copper-iron crosstalk in Arabidopsis.

Iron (Fe) and copper (Cu) are essential yet potentially toxic metals with interconnected metabolic pathways; however, the mechanisms underlying Fe-Cu crosstalk remain poorly defined. Here, we show that CITF1 (COPPER DEFICIENCY INDUCED TRANSCRIPTION FACTOR 1), a Cu homeostasis regulator in Arabidopsis thaliana, physically interacts with FIT (FER-LIKE IRON DEFICIENCY-INDUCED TRANSCRIPTION FACTOR), the central Fe homeostasis regulator, forming a nutrient-responsive transcriptional module. Under Cu deficiency, the CITF1-FIT complex accumulates and promotes expression of the Cu uptake genes COPT2 (COPPER TRANSPORTER 2), FRO4 (FERRIC REDUCTION OXIDASE 4), and FRO5 (FERRIC REDUCTION OXIDASE 5). Proteasome-dependent degradation regulates CITF1 and FIT stability, with Cu deficiency delaying their turnover in a CITF1-dependent manner. Under Fe deficiency, CITF1 expression is downregulated, allowing FIT to interact with bHLH38/39/100/101 partners and activate Fe uptake genes, as CITF1 disrupts these interactions. Thus, CITF1 negatively regulates Fe acquisition. Consistent with this, citf1-1 and citf1-2 mutants show reduced sensitivity to Fe deficiency. Under Cu deficiency, the citf1-2 and fit-2 mutants have additive effects and under Fe deficiency, the double mutant shows partial suppression of the fit-2 slow growth phenotype, supporting the positive and negative roles of CITF1 in Cu and Fe homeostasis, respectively. Complete loss of CITF1 function in the homozygous citf1-1 fit-2 double mutant causes embryo lethality, revealing roles for CITF1 and FIT in embryo development. These findings establish CITF1 as a nutrient-responsive regulator of Cu/Fe crosstalk, functioning through interactions with FIT to prioritize Cu or Fe acquisition and balance micronutrient homeostasis.