RNA silencing of mitochondrial m-Nfs1 reduces Fe-S enzyme activity both in mitochondria and cytosol of mammalian cells.
In prokaryotes and yeast, the general mechanism of biogenesis of iron-sulfur (Fe-S) clusters involves activities of several proteins among which IscS and Nfs1p provide, through cysteine desulfuration, elemental sulfide for Fe-S core formation. Although these proteins have been well characterized, the role of their mammalian homolog in Fe-S cluster biogenesis has never been evaluated. We report here the first functional study that implicates the putative cysteine desulfurase m-Nfs1 in the biogenesis of both mitochondrial and cytosolic mammalian Fe-S proteins. Depletion of m-Nfs1 in cultured fibroblasts through small interfering RNA-based gene silencing significantly inhibited the activities of mitochondrial NADH-ubiquinone oxidoreductase (complex I) and succinate-ubiquinone oxidoreductase (complex II) of the respiratory chain, as well as aconitase of the Krebs cycle, with no alteration in their protein levels. Activity of cytosolic xanthine oxidase, which holds a [2Fe-2S] cluster, was also specifically reduced, and iron-regulatory protein-1 was converted from its [4Fe-4S] aconitase form to its apo- or RNA-binding form. Reduction of Fe-S enzyme activities occurred earlier and more markedly in the cytosol than in mitochondria, suggesting that there is a mechanism that primarily dedicates m-Nfs1 to the biogenesis of mitochondrial Fe-S clusters in order to maintain cell survival. Finally, depletion of m-Nfs1, which conferred on apo-IRP-1 a high affinity for ferritin mRNA, was associated with the down-regulation of the iron storage protein ferritin.
Fosset
Cédric
C
Institut de Chimie des Substances Naturelles, Centre National de la Recherche Scientifique, Avenue de la Terrasse, 91190 Gif-sur-Yvette, France.
Chauveau
Marie-Jeanne
MJ
Guillon
Blanche
B
Canal
Frédéric
F
Drapier
Jean-Claude
JC
Bouton
Cécile
C
D000818:Animals; D013437:Carbon-Sulfur Lyases / Q000737:chemistry / Q000235:genetics / Q000502:physiology*; D003600:Cytosol / Q000378:metabolism*; D015536:Down-Regulation; D042967:Electron Transport Complex I / Q000737:chemistry; D042963:Electron Transport Complex II / Q000737:chemistry; D005293:Ferritins / Q000737:chemistry; D007506:Iron-Sulfur Proteins / Q000737:chemistry*; D051379:Mice; D008928:Mitochondria / Q000201:enzymology / Q000378:metabolism*; D041681:NIH 3T3 Cells; D034622:RNA Interference; D014969:Xanthine Oxidase / Q000737:chemistry
D007506:Iron-Sulfur Proteins; D005293:Ferritins; D014969:Xanthine Oxidase; D042963:Electron Transport Complex II; D013437:Carbon-Sulfur Lyases; C513493:Nfs1 protein, mouse; C080546:cysteine desulfurase; D042967:Electron Transport Complex I