Deletion of the Mitochondrial Carrier Genes MRS3 andMRS4 Suppresses Mitochondrial Iron Accumulation in a Yeast Frataxin-deficient Strain

The mitochondrial solute carriers Mrs3p and Mrs4p were originally isolated as multicopy suppressors of intron splicing defects. We show here that MRS4 is co-regulated with the iron regulon genes, and up-regulated in a strain deficient for Yfh1p, the yeast homologue of human frataxin. Using in vivo 55Fe cell radiolabeling we show that in glucose-grown cells mitochondrial iron accumulation is 5–15 times higher in ΔYFH1 than in wild-type strain. However, although in a ΔYFH1ΔMRS3ΔMRS4 strain, the intracellular 55Fe content is extremely high, the mitochondrial iron concentration is decreased to almost wild-type levels. Moreover, ΔYFH1ΔMRS3ΔMRS4 cells grown in high iron media do not lose their mitochondrial genome. Conversely, a ΔYFH1 strain overexpressing MRS4 has an increased mitochondrial iron content and no mitochondrial genome. Therefore, MRS4 is required for mitochondrial iron accumulation in ΔYFH1 cells. Expression of the iron regulon and intracellular 55Fe content are higher in a ΔMRS3ΔMRS4 strain than in the wild type. Nevertheless, the mitochondrial 55Fe content, a balance between iron uptake and exit, is decreased by a factor of two. Moreover, 55Fe incorporation into heme by ferrochelatase is increased in an MRS4-overexpressing strain. The function ofMRS4 in iron import into mitochondria is discussed. The mitochondrial solute carriers Mrs3p and Mrs4p were originally isolated as multicopy suppressors of intron splicing defects. We show here that MRS4 is co-regulated with the iron regulon genes, and up-regulated in a strain deficient for Yfh1p, the yeast homologue of human frataxin. Using in vivo 55Fe cell radiolabeling we show that in glucose-grown cells mitochondrial iron accumulation is 5–15 times higher in ΔYFH1 than in wild-type strain. However, although in a ΔYFH1ΔMRS3ΔMRS4 strain, the intracellular 55Fe content is extremely high, the mitochondrial iron concentration is decreased to almost wild-type levels. Moreover, ΔYFH1ΔMRS3ΔMRS4 cells grown in high iron media do not lose their mitochondrial genome. Conversely, a ΔYFH1 strain overexpressing MRS4 has an increased mitochondrial iron content and no mitochondrial genome. Therefore, MRS4 is required for mitochondrial iron accumulation in ΔYFH1 cells. Expression of the iron regulon and intracellular 55Fe content are higher in a ΔMRS3ΔMRS4 strain than in the wild type. Nevertheless, the mitochondrial 55Fe content, a balance between iron uptake and exit, is decreased by a factor of two. Moreover, 55Fe incorporation into heme by ferrochelatase is increased in an MRS4-overexpressing strain. The function ofMRS4 in iron import into mitochondria is discussed. 4′,6′-diamino-2-phenylindole Mitochondria utilize most of the cellular iron. First, the mitochondrial ferrochelatase catalyzes the insertion of ferrous iron into protoporphyrin IX, the heme precursor of cytochromes. Second, the mitochondrial matrix and respiratory chain contain several iron-sulfur proteins. Moreover, it has recently been discovered that iron-sulfur clusters are synthesized inside mitochondria by a specific machinery involving more than 10 proteins that have orthologues in bacteria (1Muhlenhoff U. Lill R. Biochim. Biophys. Acta. 2000; 1459: 370-382Crossref PubMed Scopus (181) Google Scholar,2Craig E.A. Voisine C. Schilke B. Biol. Chem. 1999; 380: 1167-1173Crossref PubMed Scopus (42) Google Scholar). These iron-sulfur clusters are used for both mitochondrial and cytosolic proteins, and their export into the cytosol is probably mediated by the ABC transporter Atm1p (3Kispal G. Csere P. Prohl C. Lill R. EMBO J. 1999; 18: 3981-3989Crossref PubMed Scopus (585) Google Scholar). Based on their capacity to restore the low iron growth defect of an erg25 mutant (4Li L. Kaplan J. J. Biol. Chem. 1997; 272: 28485-28493Abstract Full Text Full Text PDF PubMed Scopus (72) Google Scholar), two homologous transporters belonging to the cation efflux transporter family (5Paulsen I.T. Sliwinski M.K. Nelissen B. Goffeau A. Saier M.H., Jr. FEBS Lett. 1998; 430: 116-125Crossref PubMed Scopus (196) Google Scholar), Mmt1p and Mmt2p, have been reported to play a role in mitochondrial iron transport (4Li L. Kaplan J. J. Biol. Chem. 1997; 272: 28485-28493Abstract Full Text Full Text PDF PubMed Scopus (72) Google Scholar). However, the double deletion strain does not exhibit any respiratory defect, suggesting that the role played by MMT1 and MMT2 in mitochondrial iron metabolism is modest (4Li L. Kaplan J. J. Biol. Chem. 1997; 272: 28485-28493Abstract Full Text Full Text PDF PubMed Scopus (72) Google Scholar, 6Lange H. Kispal G. Lill R. J. Biol. Chem. 1999; 274: 18989-18996Abstract Full Text Full Text PDF PubMed Scopus (133) Google Scholar). Using DNA microarrays, we have shown that mitochondria play a key role in cellular iron homeostasis (7Foury F. Talibi D. J. Biol. Chem. 2001; 276: 7762-7768Abstract Full Text Full Text PDF PubMed Scopus (161) Google Scholar). This conclusion was reached by using a strain that has been deleted for YFH1, the yeast frataxin gene (8Babcock M. de Silva D. Oaks R. Davis-Kaplan S. Jiralerspong S. Montermini L. Pandolfo M. Kaplan J. Science. 1997; 276: 1709-1712Crossref PubMed Scopus (817) Google Scholar, 9Foury F. Cazzalini O. FEBS Lett. 1997; 411: 373-377Crossref PubMed Scopus (342) Google Scholar). Frataxin is a small hydrophilic protein of unknown function conserved in the mitochondria of all eukaryotes. It has been reported that a multimeric form of frataxin is involved in iron sequestration (10Adamec J. Rusnak F. Owen W.G. Naylor S. Benson L.M. Gacy A.M. Isaya G. Am. J. Hum. Genet. 2000; 67: 549-562Abstract Full Text Full Text PDF PubMed Scopus (229) Google Scholar). In glucose-grown cells YFH1 deletion elicits mitochondrial iron overload (8Babcock M. de Silva D. Oaks R. Davis-Kaplan S. Jiralerspong S. Montermini L. Pandolfo M. Kaplan J. Science. 1997; 276: 1709-1712Crossref PubMed Scopus (817) Google Scholar, 9Foury F. Cazzalini O. FEBS Lett. 1997; 411: 373-377Crossref PubMed Scopus (342) Google Scholar) and prevents iron export from mitochondria (11Radisky D.C. Babcock M.C. Kaplan J. J. Biol. Chem. 1999; 274: 4497-4499Abstract Full Text Full Text PDF PubMed Scopus (248) Google Scholar). This phenotype is associated with increased expression of the genes involved in iron mobilization in anAFT1-dependent manner (7Foury F. Talibi D. J. Biol. Chem. 2001; 276: 7762-7768Abstract Full Text Full Text PDF PubMed Scopus (161) Google Scholar). Aft1p is an iron-sensing transcription factor that activates the transcription of a set of genes, the “iron regulon,” under cellular iron starvation conditions (12Yamaguchi-Iwai Y. Dancis A. Klausner R.D. EMBO J. 1995; 14: 1231-1239Crossref PubMed Scopus (313) Google Scholar). By analyzing published data of gene expression profiles obtained under 300 different conditions (13Hughes T.R. Marton M.J. Jones A.R. Roberts C.J. Stoughton R. Armour C.D. Bennett HA Coffey E. Dai H., He, Y.D. Kidd M.J. King A.M. Meyer M.R. Slade D. Lum P.Y. Stepaniants S.B. Shoemaker D.D. Gachotte D. Chakraburtty K. Simon J. Bard M. Friend S.H. Cell. 2000; 102: 109-126Abstract Full Text Full Text PDF PubMed Scopus (2090) Google Scholar) we found that theMRS4 and MMT2 genes are co-regulated with severalAFT1-dependent genes. These data suggest thatMRS4 and MMT2 play a role in iron metabolism. The homologous Mrs3p and Mrs4p proteins are located in the mitochondrial inner membrane and belong to the mitochondrial solute carrier family (14Wiesenberger G. Link T.A. von Ahsen U. Waldherr M. Schweyen R.J. J. Mol. Biol. 1991; 217: 23-37Crossref PubMed Scopus (78) Google Scholar). Saccharomyces cerevisiae contains 35 of these carriers (15el Moualij B. Duyckaerts C. Lamotte-Brasseur J. Sluse F.E. Yeast. 1997; 13: 573-581Crossref PubMed Scopus (93) Google Scholar, 16Belenkiy R. Haefele A. Eisen M.B. Wohlrab H. Biochim. Biophys. Acta. 2000; 1467: 207-218Crossref PubMed Scopus (43) Google Scholar), the most studied of which is the mitochondrial ATP/ADP translocator. Mitochondrial carriers are characterized by three homologous domains, each containing two transmembrane spans.MRS3 and MRS4 were initially isolated as high copy number suppressors of mitochondrial mRNA splicing defects, particularly in mrs2 mutants (17Waldherr M. Ragnini A. Jank B. Teply R. Wiesenberger G. Schweyen R.J. Curr. Genet. 1993; 24: 301-306Crossref PubMed Scopus (60) Google Scholar). MRS2 encodes a mitochondrial inner membrane protein belonging to the bacterial CORA family of magnesium transporters and is involved in magnesium homeostasis (18Bui D.M. Gregan J. Jarosch E. Ragnini A. Schweyen R.J.J. Biol. Chem. 1999; 274: 20438-20443Abstract Full Text Full Text PDF Scopus (142) Google Scholar). Mrs3p and Mrs4p carriers have been conserved during evolution, and their human homologue can rescue the thermosensitive growth defect observed in a double ΔMRS3ΔMRS4 deletion yeast strain (19Li F.Y. Nikali K. Gregan J. Leibiger I. Leibiger B. Schweyen R. Larsson C. Suomalainen A. FEBS Lett. 2001; 494: 79-84Crossref PubMed Scopus (38) Google Scholar). It has been proposed that these carriers play a role in metal transport. We thus decided to investigate whether MRS3 andMRS4 genes play a role in mitochondrial iron import. S. cerevisiae strains used in this study were the wild-type strain W303–1B (MAT α ura3-52, leu2-3, 112, trp1-1, his3-11,15, ade2-1) and (MAT α ura3-52, leu2-3, 112, trp1-1, his3-11,15, F. Cazzalini O. FEBS Lett. 1997; 411: 373-377Crossref PubMed Scopus (342) Google Scholar). (MAT α ura3-52, leu2-3, 112, trp1-1, his3-11,15, is a of a a deleted copy The ΔMRS3ΔMRS4 and deletion strains were obtained by of W303–1B with the gene U. S. J. 24: PubMed Scopus Google Scholar). This contains the and can by homologous between yeast with a containing the under the of the U. S. J. 24: PubMed Scopus Google Scholar). First, yeast strains were by the on each by homologous to the and of the gene of A. A. R. P. Yeast. 13: Scopus Google Scholar). The were as and and deleted strains were thus the deletion was by the and a deletion obtained by chain of the DNA of deleted strain was used to the deleted strain and a double deletion strain. and were The mutants and ΔYFH1ΔMRS3ΔMRS4 are from between and In the ΔYFH1ΔMRS3ΔMRS4 strain, theMRS4 deletion was obtained by of a deletion containing the by DNA to the and of the MRS4 The strains were grown in yeast in containing yeast with required and contains yeast yeast and is with an in the cells were to a concentration of in of the containing and with for were and two times with a of and 10 of and mitochondria was as F. Cazzalini O. FEBS Lett. 1997; 411: 373-377Crossref PubMed Scopus (342) Google Scholar). The of 55Fe was by of the in the mitochondrial and The import into isolated mitochondria by the of was as H. Kispal G. Lill R. J. Biol. Chem. 1999; 274: 18989-18996Abstract Full Text Full Text PDF PubMed Scopus (133) Google Scholar). the of the of their (13Hughes T.R. Marton M.J. Jones A.R. Roberts C.J. Stoughton R. Armour C.D. Bennett HA Coffey E. Dai H., He, Y.D. Kidd M.J. King A.M. Meyer M.R. Slade D. Lum P.Y. Stepaniants S.B. Shoemaker D.D. Gachotte D. Chakraburtty K. Simon J. Bard M. Friend S.H. Cell. 2000; 102: 109-126Abstract Full Text Full Text PDF PubMed Scopus (2090) Google Scholar), (13Hughes T.R. Marton M.J. Jones A.R. Roberts C.J. Stoughton R. Armour C.D. Bennett HA Coffey E. Dai H., He, Y.D. Kidd M.J. King A.M. Meyer M.R. Slade D. Lum P.Y. Stepaniants S.B. Shoemaker D.D. Gachotte D. Chakraburtty K. Simon J. Bard M. Friend S.H. Cell. 2000; 102: 109-126Abstract Full Text Full Text PDF PubMed Scopus (2090) Google Scholar) a data in which for the genes most with a gene with a The number of data used in the and the number of times the gene was a were of cells grown in the of growth was with in and in were and concentration was as reported J. PubMed Scopus Google Scholar). and were as (7Foury F. Talibi D. J. Biol. Chem. 2001; 276: 7762-7768Abstract Full Text Full Text PDF PubMed Scopus (161) Google Scholar). and were by on with for genes co-regulated with iron we a in the data by (13Hughes T.R. Marton M.J. Jones A.R. Roberts C.J. Stoughton R. Armour C.D. Bennett HA Coffey E. Dai H., He, Y.D. Kidd M.J. King A.M. Meyer M.R. Slade D. Lum P.Y. Stepaniants S.B. Shoemaker D.D. Gachotte D. Chakraburtty K. Simon J. Bard M. Friend S.H. Cell. 2000; 102: 109-126Abstract Full Text Full Text PDF PubMed Scopus (2090) Google Scholar). Using DNA microarrays, the have obtained expression profiles for the of S. cerevisiae under 300 different for the genes most with iron regulon genes as to ofMRS4 and MMT2 genes. a for genes with MRS4 iron regulon genes. We have shown that the iron regulon is up-regulated in a ΔYFH1 strain (7Foury F. Talibi D. J. Biol. Chem. 2001; 276: 7762-7768Abstract Full Text Full Text PDF PubMed Scopus (161) Google Scholar). Therefore, we the expression ofMRS4 and MMT2 in wild-type and ΔYFH1 strains grown in By we found that the mRNA of MRS4 and MMT2 were times increased in a ΔYFH1 strain It expression was low with In with iron regulon genes as O. J. J. D. C. J. Biol. Chem. 2001; 276: Full Text Full Text PDF PubMed Scopus (133) Google was in strains and a higher than in DNA have shown that is by iron a of E. J. Biol. Chem. 2001; 276: Full Text Full Text PDF PubMed Scopus (161) Google Scholar, S. E. U. S. A. 2001; PubMed Scopus Google Scholar). and deletion strains were for and MMT2 genes in wild-type and ΔYFH1 strains were However, cellular growth was decreased both in and media The growth on not from of mutants This cell metabolism not inside mitochondria ΔMRS3ΔMRS4 cells in low iron media containing an iron Moreover, growth of ΔMRS3ΔMRS4 and ΔYFH1ΔMRS3ΔMRS4 cells on was obtained of iron in the and The high iron concentration used here the growth of ΔYFH1 and the ΔMRS3ΔMRS4 deletion the iron the ΔYFH1 strain. In of ΔYFH1 cells with multicopy these cells were extremely and their mitochondrial genome. of the mitochondrial DNA was shown by and in with a strain of which no of growth on not These data suggest a between mitochondrial iron metabolism and growth defect was observed in the strain. iron in ΔYFH1 cells The deletion strain was not and phenotype was to that of ΔMRS3ΔMRS4 strains not of mitochondrial and strains grown in were in and and on for 10 these mitochondrial DNA is The mitochondrial DNA are in wild-type and ΔYFH1 are the can ΔYFH1 cells. was used with a a and an MRS4 and MMT2 genes play a role in the transport of iron into their deletion mitochondrial iron and mutant cells were with 55Fe for and the associated with mitochondria and was The iron that in the mitochondria of ΔYFH1 cells grown in (11Radisky D.C. Babcock M.C. Kaplan J. J. Biol. Chem. 1999; 274: 4497-4499Abstract Full Text Full Text PDF PubMed Scopus (248) Google Scholar). Therefore, in iron import into mitochondria more in a ΔYFH1 strain than in a strain in which the of the balance between uptake and efflux of mitochondrial iron is Mitochondria of glucose-grown ΔYFH1 cells of the cellular 55Fe with for wild-type mitochondria In the in ΔYFH1ΔMRS3ΔMRS4 mitochondria was to the found in wild-type mitochondria These were cells were in high iron media these the content was in wild-type and ΔYFH1 However, in the ΔYFH1 strain iron in mitochondria to the of the In in the ΔYFH1ΔMRS3ΔMRS4 strain iron in a Conversely, in a ΔYFH1 strain the content was times higher than in a ΔYFH1 strain The mitochondrial 55Fe content of ΔMRS3ΔMRS4 cells grown in was of that in the wild-type and It that although intracellular iron concentration was increased in ΔMRS3ΔMRS4 cells with iron was not to mitochondria and a the of the mitochondrial to intracellular iron content decreased from in the wild-type to in the ΔMRS3ΔMRS4 strain grown in We used isolated mitochondria to the incorporation of 55Fe into protoporphyrin by mitochondrial ferrochelatase in wild-type and MRS4-overexpressing This the of iron into mitochondria that can used by the was in the heme cells these show that of the Mrs3p and Mrs4p carriers is a factor for iron import into in ΔYFH1 cells grown in high iron media However, a of iron is into mitochondria in the of Therefore, transport iron iron uptake that have to are and MMT2 no on mitochondrial iron incorporation and accumulation and However, in ΔYFH1 cells decreased the in mitochondria suggesting that does not import iron into mitochondrial DNA is in ΔYFH1 strain F. Cazzalini O. FEBS Lett. 1997; 411: 373-377Crossref PubMed Scopus (342) Google Scholar). However, high iron accumulation of cells of mitochondrial DNA F. Cazzalini O. FEBS Lett. 1997; 411: 373-377Crossref PubMed Scopus (342) Google Scholar). of mitochondrial DNA was observed in ΔYFH1ΔMRS3ΔMRS4 cells in the of high iron in with the that iron does not in ΔYFH1ΔMRS3ΔMRS4 In the of in a ΔYFH1 strain in mitochondrial DNA cells were in the of high iron Using we have found that mitochondrial DNA in mitochondria not from increased expression of the MRS4 gene not was not in cells. We found that the expression of genes as was increased in a ΔMRS3ΔMRS4 strain, although to a than in a ΔYFH1 strain The expression of the iron regulon genes was higher in a ΔYFH1ΔMRS3ΔMRS4 strain than in a ΔYFH1 strain not These data show that the of is associated with of the that Aft1p low iron However, the of 55Fe in the of ΔMRS3ΔMRS4 strains is high that this iron is not by in a specific as the F. E. FEBS Lett. PubMed Scopus Google Scholar, L. D. Kaplan J. J. Biol. Chem. 2001; 276: Full Text Full Text PDF PubMed Scopus Google Scholar). The expression genes was not increased in cells. and strains were grown in and their content as as their and were The content was not in ΔMRS3ΔMRS4 cells and in isolated mitochondria were decreased Therefore, the of mitochondrial iron by the import was to a of heme and iron-sulfur in these cells a In ΔYFH1 the content was and was than in wild-type although the concentration by was not The in content and in and was more and MRS4 were in a ΔYFH1 strain and Therefore, in ΔYFH1ΔMRS3ΔMRS4 cells the in mitochondrial iron concentration a This that a strain that has no frataxin more mitochondrial iron than a wild-type strain for heme and iron-sulfur The mitochondrial was in all strains suggesting that proteins were The deletion of MMT1 genes no on and heme content not in ΔMRS3ΔMRS4 and cells. were in mitochondria of cells. and have been to wild-type is as the of to were using cell and mitochondrial In the of wild-type and ΔYFH1 are MRS3 and MRS4 are multicopy suppressors of mrs2 mutants (17Waldherr M. Ragnini A. Jank B. Teply R. Wiesenberger G. Schweyen R.J. Curr. Genet. 1993; 24: 301-306Crossref PubMed Scopus (60) Google Scholar). The have in magnesium transport into mitochondria (18Bui D.M. Gregan J. Jarosch E. Ragnini A. Schweyen R.J.J. Biol. Chem. 1999; 274: 20438-20443Abstract Full Text Full Text PDF Scopus (142) Google Scholar), and it has recently been shown that mitochondrial magnesium are to wild-type in an mrs2 mutant MRS4 J. M. Schweyen R.J. 2001; PubMed Scopus Google Scholar). We not in the mitochondrial magnesium concentration of strains deleted for We the of on cellular The most was obtained with of wild-type cells was in the of was for ΔYFH1 cells and for cells and In all strains of cellular growth by was associated with of mutants In ΔMRS3ΔMRS4 cells were extremely to and no of mitochondrial DNA was observed in ΔYFH1ΔMRS3ΔMRS4 cells. MRS4 on a multicopy increased to and We were not to inside mitochondria of wild-type mutant strains grown in the of However, was in mitochondria overexpressing Mrs4p accumulation in these the ΔMRS3ΔMRS4 strain a higher intracellular concentration than the wild-type strain this is not it in the were associated with increased cellular to and as and to a and magnesium in mitochondrial and of wild-type and mutant protein were in in the of Mitochondrial and of the cells were for and magnesium were under is the wild-type strain the multicopy not in a were in in the of Mitochondrial and of the cells were for and magnesium were under is the wild-type strain the multicopy not the of the capacity strains to the mitochondrial magnesium transport defect of mrs2 it has been proposed that these mitochondrial carriers transport The that MRS4 expression was co-regulated with genes involved in iron mobilization from the and intracellular to investigate whether Mrs4p iron import into The expression of the iron regulon and intracellular iron content are increased in a ΔMRS3ΔMRS4 strain. Nevertheless, the of iron in mitochondria is in a ΔMRS3ΔMRS4 Therefore, the iron from the by ΔMRS3ΔMRS4 cells is not by Aft1p and does not It in the which is an iron in yeast S. E. U. S. A. 2001; PubMed Scopus Google Scholar, F. E. FEBS Lett. PubMed Scopus Google Scholar). Mitochondrial iron content is a balance between uptake and exit, and decreased mitochondrial iron uptake in the ΔMRS3ΔMRS4 strain by that the mitochondrial iron content is Moreover, the growth of ΔMRS3ΔMRS4 cells to mitochondrial iron concentration as as data that for of the mitochondrial iron The in the incorporation of iron into protoporphyrin by ferrochelatase in isolated mitochondria overexpressing Mrs4p in a more that Mrs4p a role in mitochondrial iron transport in the wild-type strain. The is different for ΔYFH1 cells in The iron regulon is up-regulated cells are grown in high iron and a of the iron into the cell is to Moreover, no iron is from mitochondria (11Radisky D.C. Babcock M.C. Kaplan J. J. Biol. Chem. 1999; 274: 4497-4499Abstract Full Text Full Text PDF PubMed Scopus (248) Google Scholar). Therefore, the of mitochondrial iron uptake is more in a ΔYFH1 strain. the intracellular iron is increased in a ΔYFH1ΔMRS3ΔMRS4 strain grown in the of the mitochondrial iron content is times than in a ΔYFH1 strain. Moreover, although cells in high iron media lose mitochondrial this is by These data show that iron import into mitochondria is in ΔYFH1ΔMRS3ΔMRS4 cells. Conversely, MRS4 in ΔYFH1 cells elicits a in the mitochondrial iron These cells have no mitochondrial DNA under conditions and probably as a of mitochondrial iron Therefore, in ΔYFH1 cells carriers to mitochondrial iron of the MRS4 in the ΔYFH1 strain probably to the show that Mrs4p a role in mitochondrial iron This function is with transporter that has not been This unknown transporter have high for mitochondrial iron are increased in ΔYFH1ΔMRS3ΔMRS4 cells grown in high iron Mrs3p and Mrs4p low expression of the iron regulon in ΔMRS3ΔMRS4 cells is associated with increased to in and It has been shown that iron starvation is associated with increased expression of metal transporters in the membrane L. Kaplan J. J. Biol. Chem. 1998; Full Text Full Text PDF PubMed Scopus Google Scholar). of ΔMRS3ΔMRS4 cells by the increased by metal intracellular However, ΔMRS3ΔMRS4 cells exhibit an to The of mitochondrial which is observed in wild-type and ΔYFH1 strains with high is by these cells more than wild type. This that is in the the expression of which encodes a transporter is increased in ΔMRS3ΔMRS4 cells L. Kaplan J. J. Biol. Chem. 1998; Full Text Full Text PDF PubMed Scopus Google Scholar). Mitochondrial concentration was under in ΔYFH1 cells. However, was in mitochondria of MRS4-overexpressing cells. The exhibit increased to a role in yeast and more in are extremely high, and from between iron and in of as ferrochelatase H. J. Mol. Biol. PubMed Scopus Google Scholar, U. S. A. 1999; PubMed Scopus Google Scholar). It has been found that Mrs4p can for a mitochondrial inner membrane protein involved in magnesium transport (18Bui D.M. Gregan J. Jarosch E. Ragnini A. Schweyen R.J.J. Biol. Chem. 1999; 274: 20438-20443Abstract Full Text Full Text PDF Scopus (142) Google Scholar), suggesting that Mrs4p can used for import of magnesium into mitochondria J. M. Schweyen R.J. 2001; PubMed Scopus Google Scholar). transporters in the inner mitochondrial membrane that belong to the family of cation efflux Mmt1p and Mmt2p, have been reported to play a role in mitochondrial iron import (4Li L. Kaplan J. J. Biol. Chem. 1997; 272: 28485-28493Abstract Full Text Full Text PDF PubMed Scopus (72) Google Scholar). Moreover, MMT2 is co-regulated with several iron regulon genes. MMT1 and MMT2 were isolated as multicopy suppressors of the growth defect of mutant in low iron media (4Li L. Kaplan J. J. Biol. Chem. 1997; 272: 28485-28493Abstract Full Text Full Text PDF PubMed Scopus (72) Google Scholar). The found that the double deletion mutant grown in low iron media a for a cell Moreover, cells MMT2 in high iron media have increased of iron both in cytosol and We found that MMT2 the intracellular iron content, we not in the mitochondrial iron It that iron to of iron on yeast H. Kispal G. Lill R. J. Biol. Chem. 1999; 274: 18989-18996Abstract Full Text Full Text PDF PubMed Scopus (133) Google Scholar). Moreover, as reported by H. Kispal G. Lill R. J. Biol. Chem. 1999; 274: 18989-18996Abstract Full Text Full Text PDF PubMed Scopus (133) Google Scholar), we not a phenotype to iron in deletion This by the low expression of these In to a (4Li L. Kaplan J. J. Biol. Chem. 1997; 272: 28485-28493Abstract Full Text Full Text PDF PubMed Scopus (72) Google Scholar), deletion strains were to the in the mitochondrial 55Fe observed in ΔYFH1 cells overexpressing MMT2 does not the that iron into mitochondria with that MMT2 as a multicopy of the iron ΔYFH1 cells. These data suggest that Mmt1p and play a role in the efflux of iron from It has been that an in by mitochondrial iron overload is a of the mitochondrial in cells (11Radisky D.C. Babcock M.C. Kaplan J. J. Biol. Chem. 1999; 274: 4497-4499Abstract Full Text Full Text PDF PubMed Scopus (248) Google Scholar). This is probably in ΔYFH1 cells in the of high iron Mrs4p is We found that ΔYFH1 cells that do not the mitochondrial are extremely these the mitochondrial is However, in ΔYFH1 is no mitochondrial iron and a specific defect in the of iron-sulfur proteins as is Moreover, a in mitochondrial iron concentration is associated with a in This that in a strain more iron is required to iron-sulfur that mitochondrial iron is not This conclusion is in with the that of yeast frataxin of iron in a form (10Adamec J. Rusnak F. Owen W.G. Naylor S. Benson L.M. Gacy A.M. Isaya G. Am. J. Hum. Genet. 2000; 67: 549-562Abstract Full Text Full Text PDF PubMed Scopus (229) Google Scholar), suggesting that frataxin the of iron for the of iron-sulfur role for frataxin in iron-sulfur metabolism is by data obtained with that a in the of iron-sulfur proteins to mitochondrial iron H. Simon D. M. P. F. J. C. R. P. M. Genet. 2001; PubMed Scopus Google Scholar). Moreover, on the of on it has recently been proposed that frataxin a role in iron-sulfur B. P. Hum. Mol. Genet. 2001; PubMed Scopus Google Scholar). We thus that two in frataxin deficient cells. First, of iron-sulfur clusters is F. FEBS Lett. 1999; PubMed Scopus Google Scholar), and under conditions of mitochondrial iron. the We Lill for the of the F. the and for a as in the of in the of M. is for and