Abstract
Biogenesis of the iron-sulfur (Fe-S) cluster is an indispensable process in living cells. In mammalian mitochondria, the initial step of the Fe-S cluster assembly process is assisted by the NFS1-ISD11 complex, which delivers sulfur to scaffold protein ISCU during Fe-S cluster synthesis. Although ISD11 is an essential protein, its cellular role in Fe-S cluster biogenesis is still not defined. Our study maps the important ISD11 amino acid residues belonging to putative helix 1 (Phe-40), helix 3 (Leu-63, Arg-68, Gln-69, Ile-72, Tyr-76), and C-terminal segment (Leu-81, Glu-84) are critical for in vivo Fe-S cluster biogenesis. Importantly, mutation of these conserved ISD11 residues into alanine leads to its compromised interaction with NFS1, resulting in reduced stability and enhanced aggregation of NFS1 in the mitochondria. Due to altered interaction with ISD11 mutants, the levels of NFS1 and Isu1 were significantly depleted, which affects Fe-S cluster biosynthesis, leading to reduced electron transport chain complex (ETC) activity and mitochondrial respiration. In humans, a clinically relevant ISD11 mutation (R68L) has been associated in the development of a mitochondrial genetic disorder, COXPD19. Our findings highlight that the ISD11 R68A/R68L mutation display reduced affinity to form a stable subcomplex with NFS1, and thereby fails to prevent NFS1 aggregation resulting in impairment of the Fe-S cluster biogenesis. The prime affected machinery is the ETC complex, which showed compromised redox properties, causing diminished mitochondrial respiration. Furthermore, the R68L ISD11 mutant displayed accumulation of mitochondrial iron and reactive oxygen species, leading to mitochondrial dysfunction, which correlates with the phenotype observed in COXPD19 patients.
Highlights
NFS1-ISD11 complex is essential for the Fe-S cluster assembly process and the homozygous residue at position 68 into leucine (R68L) ISD11 mutation causes the mitochondrial disorder, combined oxidative phosphorylation deficiency 19 (COXPD19)
We have delineated the importance of the ISD11 protein as a part of the sulfur-transfer protein complex, NFS1-ISD11 in Fe-S cluster biogenesis pathway
Our results indicate that the ISD11 protein interacts with the cysteine desulfurase protein NFS1 and prevents its self-aggregation, assisting a functional Fe-S cluster biogenesis process in the mitochondrial compartment
Summary
NFS1-ISD11 complex is essential for the Fe-S cluster assembly process and the homozygous R68L ISD11 mutation causes the mitochondrial disorder, COXPD19. Our study maps the important ISD11 amino acid residues belonging to putative helix 1 (Phe-40), helix 3 (Leu-63, Arg-68, Gln-69, Ile-72, Tyr-76), and C-terminal segment (Leu, Glu-84) are critical for in vivo Fe-S cluster biogenesis Mutation of these conserved ISD11 residues into alanine leads to its compromised interaction with NFS1, resulting in reduced stability and enhanced aggregation of NFS1 in the mitochondria. Role of ISD11 Protein in NFS1-ISD11 Subcomplex Stability bacteria and Isu1/2 in yeast) acts as a scaffold for the assembly of Fe-S cluster before its transfer to apoproteins. We have mapped key residues on the ISD11 protein that are essential for NFS1 interaction to maintain its levels by forming a stable subcomplex that is critical for Fe-S cluster biogenesis. Our findings highlight that the R68L mutation results in impaired Fe-S cluster biogenesis, elevated mitochondrial iron, and oxidative stress, which contribute significantly toward the development of COXPD19
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