SUMMARY Redox homeostasis and Fe–S protein maturation are critical for chloroplast function. Chloroplast‐localized glutaredoxins (Grxs) are versatile enzymes that function either as oxidoreductases or Fe–S cluster transferases. However, their target proteins and underlying molecular mechanisms remain incompletely understood. Here, we demonstrate that the cysteine 34 within the active site motif of chloroplast‐localized Arabidopsis GrxS12 is the key residue governing its oxidoreductase activity, while the C‐terminal cysteine 92 plays a secondary role. These cysteines undergo glutathionylation or form an intramolecular disulfide bond, which can be reduced by Trx‐m1. Although GrxS12 is unable to bind Fe–S clusters itself, it interacts with and reduces the Fe–S cluster assembly protein SufB, thereby regulating Fe–S protein maturation. Loss of GrxS12 induces redox alterations of multiple chloroplast proteins, increased oxidation of SufB, and decreased abundances of several Fe–S proteins. These changes likely underlie the observed structural damage to chloroplasts, reduced photosynthetic efficiency, and slower growth in grxs12 mutants. Our results reveal a novel mechanism by which GrxS12 regulates chloroplast Fe–S cluster biogenesis through its oxidoreductase activity.
Liu et al. (Thu,) studied this question.