Functional F1-ATPase Essential in Maintaining Growth and Membrane Potential of Human Mitochondrial DNA-depleted ρ° Cells

Abstract

F1-ATPase assembly has been studied in human ρ° cells devoid of mitochondrial DNA (mtDNA). Since, in these cells, oxidative phosphorylation cannot provide ATP, their growth relies on glycolysis. Despite the absence of the mtDNA-coded F0 subunits 6 and 8, ρ° cells possessed normal levels of F1-ATPase α and β subunits. This F1-ATPase was functional and azide- or aurovertin-sensitive but oligomycin-insensitive. In addition, aurovertin decreased cell growth in ρ° cells and also reduced their mitochondrial membrane potential, as measured by rhodamine 123 fluorescence. Therefore, a functional F1-ATPase was important to maintain the mitochondrial membrane potential and the growth of these ρ° cells. Bongkrekic acid, a specific adenine nucleotide translocator (ANT) inhibitor, also reduced ρ° cell growth and mitochondrial membrane potential. In conclusion, ρ° cells need both a functional F1-ATPase and a functional ANT to maintain their mitochondrial membrane potential, which is necessary for their growth. ATP hydrolysis catalyzed by F1 must provide ADP3− at a sufficient rate to maintain a rapid exchange with the glycolytic ATP4− by ANT, this electrogenic exchange inducing a mitochondrial membrane potential efficient enough to sustain cell growth. However, since the effects of bongkrekic acid and of aurovertin were additive, other electrogenic pumps should cooperate with this pathway. F1-ATPase assembly has been studied in human ρ° cells devoid of mitochondrial DNA (mtDNA). Since, in these cells, oxidative phosphorylation cannot provide ATP, their growth relies on glycolysis. Despite the absence of the mtDNA-coded F0 subunits 6 and 8, ρ° cells possessed normal levels of F1-ATPase α and β subunits. This F1-ATPase was functional and azide- or aurovertin-sensitive but oligomycin-insensitive. In addition, aurovertin decreased cell growth in ρ° cells and also reduced their mitochondrial membrane potential, as measured by rhodamine 123 fluorescence. Therefore, a functional F1-ATPase was important to maintain the mitochondrial membrane potential and the growth of these ρ° cells. Bongkrekic acid, a specific adenine nucleotide translocator (ANT) inhibitor, also reduced ρ° cell growth and mitochondrial membrane potential. In conclusion, ρ° cells need both a functional F1-ATPase and a functional ANT to maintain their mitochondrial membrane potential, which is necessary for their growth. ATP hydrolysis catalyzed by F1 must provide ADP3− at a sufficient rate to maintain a rapid exchange with the glycolytic ATP4− by ANT, this electrogenic exchange inducing a mitochondrial membrane potential efficient enough to sustain cell growth. However, since the effects of bongkrekic acid and of aurovertin were additive, other electrogenic pumps should cooperate with this pathway. adenine nucleotide translocator 3-(cyclohexylamino)-1-propanesulfonic acid subunit II and IV of cytochrome oxidase dicyclohexylcarbodiimide carbonyl cyanidep-trifluoromethoxyphenylhydrazone nonyl acridine orange rhodamine 123 10 mm Tris-HCl, pH 7.6, 0.1 mm NaCl oligomycin sensitivity conferring protein polymerase chain reaction. The biogenesis of mitochondrial proteins is controlled by both nuclear and mitochondrial genomes. The proteins coded by the mtDNA are subunits of enzyme complexes involved in oxidative phosphorylation. Since all these complexes also contain proteins coded by the nuclear genome, mechanisms regulating the coordinated expression and assembly of the subunits of nuclear and mitochondrial origin must exist. In yeast cells, nuclear DNA-coded components continue to be synthesized and imported into mitochondria, even when the synthesis of mtDNA-coded subunits is blocked (cf. for review, Refs. 1Tzagoloff A. Myers A.M. Annu. Rev. Biochem. 1986; 55: 249-285Crossref PubMed Scopus (327) Google Scholar and 2Attardi G. Schatz G. Annu. Rev. Cell Biol. 1988; 4: 289-333Crossref PubMed Scopus (1113) Google Scholar). The groups of Schatz and co-workers (3Suzuki C.K. Rep M. Maarten van Dijl J. Suda K. Grivell L.A. Schatz G. Trends Biochem. Sci. 1997; 22: 118-123Abstract Full Text PDF PubMed Scopus (212) Google Scholar) and Neupert (4Neupert W. Annu. Rev. Biochem. 1997; 66: 863-917Crossref PubMed Scopus (984) Google Scholar) have shown that a mitochondrial membrane potential is a key requirement for protein import into mitochondria (5Gasser S.M. Daum G. Schatz G. J. Biol. Chem. 1982; 257: 13034-13041Abstract Full Text PDF PubMed Google Scholar). In normal cells, the mitochondrial membrane potential is mainly maintained by transmembrane proton pumping occurring during electron transfer or during ATP hydrolysis catalyzed by the ATPase-ATP synthase. In ρ° cells depleted of mtDNA, these complexes cannot be functional since all complexes involved in proton pumping contain mtDNA-coded subunits (6Anderson S. Banquier A.T. Barrel B.G. de Bruijn M.H.L. Coulson A.R. Drouin J. Eperon I.C. Nierlich D.P. Rose B.A. Sanger F. Schreier P.H. Smith A.J.H. Staden R. Young I.G. Nature. 1981; 290: 457-465Crossref PubMed Scopus (7841) Google Scholar). However, proteins of nuclear origin are imported into the ρ° cell mitochondria (7Herzberg N.H. Middelkoop E. Adorf M. Dekker H.L. Van Galen M.J. Van den Berg M. Bolhuis P.A. Van den Bogert C. Eur. J. Cell Biol. 1993; 61: 400-408PubMed Google Scholar). Therefore, the mitochondrial membrane potential must be maintained by other electrogenic pumps. In yeast, it has been suggested that the adenine nucleotide translocator (ANT)1 mediates an exchange of ATP4− synthesized in the cytosol during glycolysis for ADP3− to maintain this mitochondrial membrane potential (8Kolarov J. Klingenberg M. FEBS Lett. 1974; 45: 320-323Crossref PubMed Scopus (29) Google Scholar). However, the exact mechanism that maintains this potential has not been thoroughly investigated. Since the mtDNA-coded subunits are essential components for oxidative phosphorylation, ρ° cells lacking mtDNA rely on glycolysis for their energy demand. The NADH produced during glycolysis is then reoxidized by the lactate dehydrogenase (9Martinus R.D. Garth G.P. Webster T.L. Cartwright P. Naylor D.J. Hoj P.B. Hoogenraad N.J. Eur. J. Biochem. 1996; 240: 98-103Crossref PubMed Scopus (259) Google Scholar). In addition, several reports indicate that some nuclearly coded subunits of the respiratory chain complexes are imported and partly assembled into the mitochondria of the ρ° cells (10Nijtmans L.G.J. Spelbrink J.N. Van Galen M.J.M. Zwaan M. Klement P. Van den Bogert C. Biochim. Biophys. Acta. 1995; 1265: 117-126Crossref PubMed Scopus (46) Google Scholar). The role (if any) of these partly assembled complexes is unknown. The present study reports observations concerning the role and mechanism of assembly of the mitochondrial ATPase in human mtDNA-deficient ρ° cells. The ATPase-ATP synthase (F0F1) is made of a hydrophilic portion F1 required for enzyme catalysis and connected by a stalk to the hydrophobic sector F0, which is involved in proton translocation occurring during ATP synthesis or ATP hydrolysis. (cf. Ref. 11Boyer P.D. Annu. Rev. Biochem. 1997; 66: 717-749Crossref PubMed Scopus (1639) Google Scholar, for review). In mammals, F1 contains five different subunits: α, β, γ, δ, and ε. F0 is made of the subunits a (or subunit 6), b, c (also named subunit 9 or DCCD-binding protein), d, e, f, g, and A6L (or subunit 8); the stalk contains OSCP, F6, and parts of F0 such as subunits b and d or of F1 such as γ (12Collinson I.R. van Raaj M.J. Runswick M.J. Fearnley I.M. Skehel J.M. Orris G.L. Miroux B. Walker J.E. J. Mol. Biol. 1994; 242: 408-421PubMed Google Scholar). The subunits a (or 6) and A6L (or 8) that are coded by the mtDNA are absent in human ρ° cells. Here, we show that F1 is assembled in human ρ° cells as an azide- or aurovertin-sensitive ATPase and that this ATPase activity is essential for the ρ° cell growth because it is involved in the maintenance of their mitochondrial membrane potential. Cell culture reagents were from Life Technology Inc. except uridine, which was from Sigma. The ρ° HeLa S3 cells devoid of mtDNA were prepared by ethidium bromide treatment of HeLa S3 cells (ρ+) (ATCC CCL2.2) by Dr. J. L. Vayssière. The ρ° 143B TK− cells were obtained from 143B TK− osteosarcoma (ρ+) (ATCC CRL8303) (13Morais R. Zinkewich-Péotti K. Parent M. Wang H. Babai F. Zollinger M. Cancer Res. 1994; 54: 3889-3896PubMed Google Scholar). The and ρ° cells were to by Dr. and Dr. The bongkrekic acid was a of P. The was obtained from The α and β subunits were prepared from the and M. C. Sci. S. A. PubMed Scopus Google and the oxidase subunit II and subunit IV were prepared from the and by Dr. Biochim. Biophys. Acta. 1996; PubMed Scopus Google Scholar). The was obtained from The were from and other reagents were from or Sigma. HeLa S3 cells were at in a with in with and and mm 143B cells were in the in with and mm was in cell culture for ρ° HeLa S3 cells and ρ° 143B cells of ρ° 143B cells, which are was and ρ° cell were for at with 10 of in and was on of cells to of and with the mtDNA at to and to or to and to at the of the and of the (6Anderson S. Banquier A.T. Barrel B.G. de Bruijn M.H.L. Coulson A.R. Drouin J. Eperon I.C. Nierlich D.P. Rose B.A. Sanger F. Schreier P.H. Smith A.J.H. Staden R. Young I.G. Nature. 1981; 290: 457-465Crossref PubMed Scopus (7841) Google The was as in H. C. Biochim. Biophys. Acta. 1996; PubMed Scopus Google of and to the of DNA in the cell was made by of the nuclear for the from to and a of and were on and ρ° cells as C. Mol. Biochem. 1997; PubMed Scopus Google Scholar). The of to mitochondrial was made as and with the mtDNA at to and to or to and to to or (6Anderson S. Banquier A.T. Barrel B.G. de Bruijn M.H.L. Coulson A.R. Drouin J. Eperon I.C. Nierlich D.P. Rose B.A. Sanger F. Schreier P.H. Smith A.J.H. Staden R. Young I.G. Nature. 1981; 290: 457-465Crossref PubMed Scopus (7841) Google Scholar). The were and cell was in an were with and in mm mm pH at for the cell at in of 10 mm Tris-HCl, mm mm and mm pH were by five of by of a with a of the were with the acid to of proteins of cell were on a and to a membrane at a of in a 10 pH as a transfer The were then for in the different the were for with a in and then for with at in The activity was with the to the the were for 10 and for in The were α M. C. Sci. S. A. PubMed Scopus Google Scholar) β M. C. Sci. S. A. PubMed Scopus Google Scholar) II Biochim. Biophys. Acta. 1996; PubMed Scopus Google Scholar) and IV Biochim. Biophys. Acta. 1996; PubMed Scopus Google Scholar) were and the protein were measured as for The rate of ATP hydrolysis was measured at to A. E. J. Biol. Chem. Full Text PDF PubMed Google Scholar) by the cell to of to of mm pH mm mm mm ATP, 10 of lactate of and of NADH by the mitochondrial The was in the or absence of of the mitochondrial ATPase aurovertin or to the of ATPase activity that was to the F1 or in the cell the effects of aurovertin and bongkrekic acid on the cells were into culture at a to or the culture was and the were to aurovertin bongkrekic acid or The cells were for with a culture when The cells by treatment were by The culture was of cell treatment with bongkrekic were with The were for at The was with an of The lactate in the was at by lactate dehydrogenase of Scholar). The of which is to be to the of and of the mitochondrial membrane potential, was to the of mitochondrial in ρ° and cells J.M. A. R. Eur. J. Biochem. 1994; PubMed Scopus Google Scholar). was to the mitochondrial membrane potential Cell Biol. PubMed Scopus Google Scholar). cell was into culture at a of to and to of cell were with or aurovertin 10 bongkrekic acid for to was in some to the when the mitochondrial membrane potential was The cells were with and for at with 0.1 of or at in a with or 10 The cells were then with and the was and were measured in a with at and at Cell Biol. PubMed Scopus Google Scholar). was the was not by the of bongkrekic acid, or made the of cells by and K. H. M. Biol. 1997; PubMed Scopus Google Scholar) an of the of cells in all ρ° HeLa S3 cell growth was on the of and uridine, as shown for 143B cells (13Morais R. Zinkewich-Péotti K. Parent M. Wang H. Babai F. Zollinger M. Cancer Res. 1994; 54: 3889-3896PubMed Google Scholar). The for ρ° and HeLa S3 cells were and and that of ρ° and 143B cells, and In the ρ° cells, mtDNA be by However, the cytochrome b not be by a to the be in the ρ° cell This from into the nuclear as shown H. C. S. K. PubMed Scopus Google Scholar). The ρ° cells were devoid of functional mtDNA since mitochondrial be by not The obtained of proteins to with and as the II was absent from the of ρ° cells and present in cells. the the nuclearly IV and F1-ATPase β subunits were both in and ρ° cells. The expression of IV was in ρ° in cells, that of the F1-ATPase β subunit was in both of and ρ° cells. In a it was shown that the F1-ATPase α subunit was also in all cell not The ATPase activity was in ρ° and cell the of this activity that was to the mitochondria from that from other specific to the mitochondrial ATPase were The the activity and that obtained in the of the to the mitochondrial that which to F0 Annu. Rev. Biochem. PubMed Scopus Google not the ATPase activity of ρ° cells devoid of the coded F0 subunits 6 and However, oligomycin the ATPase activity of HeLa S3 cells by and that of 143B cells by the such as 55: PubMed Scopus Google Scholar) and aurovertin J. Biol. Chem. Full Text PDF PubMed Google which to decreased the rate of ATP hydrolysis both in ρ° and cell was the efficient inhibitor, ATP hydrolysis by in and ρ° HeLa S3 cells and by in and ρ° 143B cells. In addition, to this aurovertin-sensitive F1-ATPase activity on F1 or on partly assembled sensitivity was and show that the ρ° cell ATPase activity decreased at that of cells and that studied of ρ° or cells at at or of this activity was for the ρ° HeLa S3 or ρ° 143B cells, at it decreased by for the ρ° HeLa S3 cells and was for the ρ° 143B cells. The ATPase activity was also for the 143B cells at both it decreased for the by and in HeLa S3 cells at and the functional F1-ATPase activity a role in the of ρ° cells, the effects of aurovertin were on the growth of ρ° and cells. The of cell growth of aurovertin treatment was with cells. The sensitivity to aurovertin was in in ρ° cells, since of cell was obtained at aurovertin for HeLa S3 cells and for ρ° HeLa S3 cells or and 10 for 143B cells and for ρ° 143B cells Therefore, the of the mitochondrial F1-ATPase activity decreased cell growth in ρ° cells as as in the cells, the aurovertin the was in ρ° in cells. In addition, the aurovertin necessary to the 143B cell growth was that the HeLa S3 cells. The of was measured on both ρ° and cell to their mitochondrial membrane potential. The of was measured in to the of mitochondrial that the was in ρ° and HeLa S3 cells and was in ρ° 143B in 143B cells that the mitochondrial membrane potential was of the of in ρ° HeLa S3 and HeLa S3 cells In ρ° 143B cells, it was in 143B cells, but the was important that for In the of HeLa S3 cells, a treatment with 0.1 decreased the mitochondrial membrane potential of both ρ° and cells to a In the of 143B cells, decreased mitochondrial membrane potential, but was important with ρ° 143B cells. However, the in the of a in ρ° and 143B cells. Therefore, in the absence of mitochondrial membrane potential, the obtained with the to to the of in the cells. the from to 10 not the in cell In conclusion, the mitochondrial membrane potential, as by is in ρ° and HeLa cells, that in ρ° 143B cells is 10 to that of the 143B cells. of the mitochondrial membrane potential of ρ° 143B cells, the mitochondrial membrane potential were with HeLa S3 cells. The effects of aurovertin were on the mitochondrial membrane potential of ρ° and HeLa S3 cells, as measured by fluorescence. that in and ρ° HeLa a of mitochondrial membrane potential was a were obtained with a 6 Since the cell was not a or a aurovertin the aurovertin on mitochondrial membrane potential is not to a cell Since the respiratory chain is not functional in ρ° cells, the membrane potential must be by a mechanism of this The ANT, which ATP4− the of the is to in the maintenance of this potential, as suggested for ρ° yeast cells (8Kolarov J. Klingenberg M. FEBS Lett. 1974; 45: 320-323Crossref PubMed Scopus (29) Google Scholar). the role by the F1-ATPase be the ANT, the of bongkrekic acid was on the growth and the mitochondrial membrane potential of both ρ° and cells. that bongkrekic acid the 143B cells growth that of the ρ° 143B cells of treatment even at the the an of ρ° S3 cell growth was of treatment with 10 bongkrekic the be a treatment not In the of the HeLa S3 cells, was that 10 acid the cell growth by but a culture was this was the growth was to Since, in the of bongkrekic acid, the ATP produced by oxidative phosphorylation cannot the the cell must then ATP glycolysis. In this the NADH produced during glycolysis must be and lactate should be this the lactate was measured in and ρ° HeLa S3 cells In the absence of bongkrekic acid, the lactate was 6 in ρ° in cells, as since in ρ° cells the ATP relies on glycolysis. The lactate in HeLa S3 cells with 10 bongkrekic acid, even the of cells was the the lactate decreased in ρ° HeLa S3 cells with 10 bongkrekic However, the of ρ° cells was the lactate was with the of cells, it in cells and in ρ° cells. Bongkrekic acid not the mitochondrial membrane potential of HeLa S3 cells a treatment or a treatment not the in ρ° HeLa S3 cells, the mitochondrial membrane potential was reduced a treatment as as a F1-ATPase and ANT or in to maintain the ρ° HeLa S3 cell growth and mitochondrial membrane potential, the effects of aurovertin and bongkrekic acid were The of aurovertin or 10 bongkrekic acid decreased the cell growth by a the of bongkrekic acid and aurovertin decreased it by to that of the The ρ° cell growth with aurovertin or bongkrekic acid was that with 0.1 or of of the to the effects on cell the mitochondrial membrane potential was decreased in the of aurovertin and bongkrekic acid when was aurovertin was to was on cell growth or on mitochondrial membrane potential. that ρ° cells devoid of the subunits 6 and contain an F1-ATPase essential for their growth. The and of F1 α and β subunits were in ρ° and cells. Therefore, the α and β subunit were imported in ρ° cells, and their were in the have shown that the ρ° 143B cells the of F1 β subunit as that of the the of some other mitochondrial proteins such as cytochrome cytochrome oxidase subunit IV and and ANT and ANT were K. P.D. J. 1995; PubMed Google Scholar). of mitochondrial protein synthesis in human cells also decreased the of or IV subunits of nuclear origin that of the F1-ATPase α and β subunits during culture (10Nijtmans L.G.J. Spelbrink J.N. Van Galen M.J.M. Zwaan M. Klement P. Van den Bogert C. Biochim. Biophys. Acta. 1995; 1265: 117-126Crossref PubMed Scopus (46) Google Scholar). The that respiratory chain subunits of nuclear origin were when were not In the of the assembly of has been in yeast mitochondria L.A. Eur. J. Biochem. PubMed Scopus Google Scholar). In the human ρ° cell studied the cytochrome oxidase subunit IV was decreased culture This that at some of the cytochrome oxidase subunits of nuclear origin be imported In the of the the normal of the α and β subunits suggested that F1 be assembled and The ATPase activity measured in the human ρ° cells was to this mitochondrial it was aurovertin-sensitive J. Biol. Chem. Full Text PDF PubMed Google Scholar). the since oligomycin sensitivity Annu. Rev. Biochem. PubMed Scopus Google Scholar) on subunit 6 Eur. J. Biochem. PubMed Scopus Google cells were by Therefore, F1 was assembled and in human ρ° cells the absence of subunits 6 and not with the subunits the stalk F1 to F0 since the ATPase activity was it has been shown in F1 that the of F1 to B. L. J. PubMed Scopus Google Scholar, I.R. van Raaj M.J. Runswick M.J. Fearnley I.M. Skehel J.M. Orris G.L. Miroux B. Walker J.E. J. Mol. Biol. 1994; 242: 408-421PubMed Google Scholar) or to other subunits the stalk such as subunits b, F6, and d (12Collinson I.R. van Raaj M.J. Runswick M.J. Fearnley I.M. Skehel J.M. Orris G.L. Miroux B. Walker J.E. J. Mol. Biol. 1994; 242: 408-421PubMed Google Scholar) F1 sensitivity to Therefore, the of the F1-ATPase in human ρ° cells that at some of the stalk subunits are However, F1 be to the a for F1 on F0 in the absence of and have been in the enzyme G. L. L. Biochem. Biophys. Scopus Google Scholar). assembly of the F1 subunits in a and has been in yeast ρ° cells. In addition, this F1 activity was by the of the mitochondrial membrane and F1 was J.M. S. J. PubMed Scopus Google Scholar). In yeast, the mtDNA not for the subunits 6 and as in human but also for the DCCD-binding Therefore, a F1 was in yeast, this must also be in human ρ° cells that contain the DCCD-binding the of the assembly has not been studied in mammals, have been yeast lacking or several subunits. was in yeast, subunit 6 was not essential for the of F1 subunits to components of the F0 S. Biochim. Biophys. Acta. PubMed Scopus Google Scholar). The assembly of subunits involved the of subunits 9 protein), 8, and 6 to a and other subunits b and OSCP, were not in the when the F0 sector was not assembled S. M.J. S. Biochim. Biophys. Acta. 1988; PubMed Scopus Google Scholar, S. Biochim. Biophys. Acta. PubMed Scopus Google Scholar). However, the b subunit be to a subunit J. de G. M. B. Eur. J. Biochem. PubMed Scopus Google and a of F1 to the membrane required the of the d subunit J. J. Biol. Chem. Full Text PDF PubMed Google Scholar). the assembly is for human ρ° cells, F1 should to the membrane of the d and the of F1 the of the DCCD-binding protein and that of the b OSCP, not be with the membrane since the subunit 6 is This the sensitivity of the human ρ° cell ATPase this ATPase activity was essential to maintain the human ρ° cell growth. of the F1-ATPase activity decreased ρ° cell growth. The for the 143B cell for the HeLa S3 cell the of the ATPase activity by aurovertin was of the of in both cell This is to of these of cells. cannot be in the 143B cells, some have been (cf. Ref. 1996; PubMed Scopus Google Scholar, for review). This the levels of to the the cell growth sensitivity to aurovertin or to bongkrekic acid was in 143B in HeLa S3 cells. However, aurovertin decreased cell growth in both ρ° cell The sensitivity to aurovertin was in in ρ° cells. This is to the during the ethidium bromide treatment to the into ρ° cells, the cells must have an to their ATP the glycolysis. This is by the lactate in ρ° in cells in the absence of In ρ° cells, oxidative phosphorylation cannot ATP to sustain cell and the glycolytic must be to provide the ATP necessary to the cell energy the NADH produced during glycolysis must be reoxidized for the lactate which lactate In ρ° HeLa S3 cells, the mitochondrial membrane potential, as measured by the as that of HeLa S3 cells. The of a normal mitochondrial membrane potential in other ρ° cells has been ρ° cells (9Martinus R.D. Garth G.P. Webster T.L. Cartwright P. Naylor D.J. Hoj P.B. Hoogenraad N.J. Eur. J. Biochem. 1996; 240: 98-103Crossref PubMed Scopus (259) Google Scholar) as as human ρ° HeLa cells P. G. Biochem. Biophys. Res. PubMed Scopus Google Scholar) to the as the cells. the ρ° 143B cells show which is 10 to that of 143B cells. This be to an ρ° HeLa S3 cells and ρ° 143B cells. is that the ρ° 143B cells a mitochondrial membrane potential, which be sufficient for their growth. In such a a mitochondrial membrane potential to 10 to of the normal mitochondrial membrane potential be enough to maintain cell to sustain protein import into the The mitochondrial membrane potential the ρ° 143B cells are for to the ρ° HeLa S3 cells, which are as as the cells, ρ° 143B cells cannot a cannot that the 143B cells a to during the ethidium bromide treatment to the ρ° cells from the that the mitochondrial membrane potential measured in these ρ° 143B cells be it is in reduced the ρ° HeLa S3 cell growth rate and their mitochondrial membrane potential by as it for HeLa S3 cells. The mitochondrial membrane potential, as as the growth of ρ° HeLa S3 cells was also reduced by bongkrekic the bongkrekic acid not the HeLa S3 mitochondrial membrane potential and decreased their growth the However, bongkrekic acid ANT in the cells since bongkrekic acid lactate and energy from oxidative phosphorylation to glycolysis. The for the sensitivity of the ρ° HeLa S3 to bongkrekic acid and aurovertin is in ρ° cells, the mitochondrial membrane potential was by the ANT ATP4− for ADP3− produced by an The of aurovertin reduced the rate of in the mitochondrial to a in the nucleotide exchange which to maintain the mitochondrial membrane potential necessary for an efficient cell growth. However, the of or the other a F1 was ATP be by other mitochondrial as for involved in the import of mitochondrial proteins of nuclear origin (3Suzuki C.K. Rep M. Maarten van Dijl J. Suda K. Grivell L.A. Schatz G. Trends Biochem. Sci. 1997; 22: 118-123Abstract Full Text PDF PubMed Scopus (212) Google Scholar). the ANT was other electrogenic such as that L. M. M. H. Biochem. Biophys. 1993; PubMed Scopus Google in maintain this membrane potential or not also in this mechanism A. Biochem. J. 1993; 290: PubMed Scopus Google Scholar). observations are with by and J. Eur. J. Biochem. 1997; PubMed Scopus Google Scholar) that cells lacking the F1 subunit a growth and a membrane potential with that for ρ° cells. In conclusion, an F1-ATPase is for the maintenance of the mitochondrial membrane potential essential for human ρ° cell growth. The Dr. and Dr. for the ρ° and cells and Dr. for of the cytochrome oxidase subunits II and