Evidence of direct iron delivery by ABCI10 to DJA5/6 for chloroplast Fe-S cluster biogenesis

Iron-sulfur (Fe-S) clusters are among the oldest cofactors in biology, required for electron transfer, catalysis and regulatory switching. Consequently, chloroplasts harbor numerous Fe-S protein-dependent processes, ranging from photosynthesis to metabolism. The assembly and transfer reactions of chloroplast Fe-S cluster biogenesis are increasingly well defined, yet how iron reaches the assembly machinery has remained unresolved. Previous studies have demonstrated that the DnaJ proteins DJA5 and DJA6 can bind iron and directly facilitate its incorporation into SUFBC₂D, the scaffold for Fe-S assembly in chloroplasts. Yet, the source of iron for DJA5/6 remains unknown. Here, we show that impairing the chloroplast inner membrane iron transporter PERMEASE IN CHLOROPLASTS 1 (PIC1) does not appreciably affect SUFB and SUFC content, whereas mutating ABCI10, a subunit of a prokaryotic-type energy coupling factor (ECF) transporter, reduces the abundance of SUFB and SUFC. This indicates that ABCI10 is required for the abundance of the SUFBC₂D complex, consistent with the requirement for DJA5/6 in maintaining SUFBC₂D content. Bimolecular fluorescence complementation (BiFC) and yeast two-hybrid (Y2H) assays further show that ABCI10 interacts with DJA5/6, but not with ABCI12. Taken together, we propose that ABCI10, a subunit of the prokaryotic-type ECF transporter, contributes to the iron supply required by SUFBC₂D and is spatially positioned to deliver iron to DJA5/6 for Fe-S biosynthesis in chloroplasts, whereas PIC1 is not the major iron donor for this pathway.