Biogenesis of mitochondria: Defective yeast H+-ATPase assembled in the absence of mitochondrial protein synthesis is membrane associated

Orian, Jacqueline M.; Hadikusumo, Richard G.; Marzuki, Sangkot; Linnane, Anthony W.

doi:10.1007/bf00743246

Cited by 16 publications

(7 citation statements)

References 39 publications

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“…In addition, this F1 H ϩ -ATPase activity was modulated by the insaturation degree of the mitochondrial membrane phospholipids, and therefore F1 was membrane-bound (32). In yeast, the mtDNA codes not only for the subunits 6 and 8 as in human but also for the DCCD-binding protein.…”

Section: Discussionmentioning

confidence: 99%

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

Buchet

Godinot

1998

Journal of Biological Chemistry

219

175

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F1-ATPase assembly has been studied in human °c ells 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, °cells possessed normal levels of F1-ATPase ␣ and ␤ subunits. This F1-ATPase was functional and azide-or aurovertinsensitive but oligomycin-insensitive. In addition, aurovertin decreased cell growth in °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 °c ells. Bongkrekic acid, a specific adenine nucleotide translocator (ANT) inhibitor, also reduced °cell growth and mitochondrial membrane potential. In conclusion, °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 ADP 3؊ at a sufficient rate to maintain a rapid exchange with the glycolytic ATP 4؊ 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.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. 1 and 2). The groups of Schatz and co-workers (3) and Neupert (4) have shown that a mitochondrial membrane potential is a key requirement for protein import into mitochondria (5). 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 °cells depleted of mtDNA, these complexes cannot be functional since all complexes involved in proton pumping contain mtDNA-coded subunits (6). However, proteins of nuclear origin are imported into the °cell mitochondria (7). 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 ATP 4Ϫ synthesized in the cytosol during glycolysis for ADP 3Ϫ to maintain this mitochondrial membrane potential (8). However, the exact mechanism that maintains this potential has not been thoroughly investigated. Since the mtDNA-coded subunits are essential components for ...

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Section: Discussionmentioning

confidence: 99%

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

Buchet

Godinot

1998

Journal of Biological Chemistry

219

175

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“…Previous reports have demonstrated that rho-mitochondria contain a labile oligomycin-insensitive ATPase (F,) [37], although this complex seems slightly bound to the membrane [38]. Upon hypotonic treatment, PVY6 mitochondrial ATPase activity appeared slightly more labile than that of rho-mitochondria.…”

Section: Mod$ication Of the Atp Synlhasementioning

confidence: 95%

The role of subunit 4, a nuclear‐encoded protein of the F₀ sector of yeast mitochondrial ATP synthase, in the assembly of the whole complex

Paul

Velours

Châteaubodeau

et al. 1989

European Journal of Biochemistry

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The yeast nuclear gene ATP4, encoding the ATP synthase subunit 4, was disrupted by insertion into the middle of it the selective marker URA3. Transformation of the Saccharomyces cerevisiae strain D273-10R/A/U produced a mutant unable to grow on glycerol medium. The ATP4 gene is unique since subunit 4 was not present in mutant mitochondria; the hypothetical truncated subunit 4 was never detected. ATPase was rendered oligomycininsensitive and the F, sector of this mutant appeared loosely bound to the membrane. Analysis of mitochondrially translated hydrophobic subunits of Fo revealed that subunits 8 and 9 were present, unlike subunit 6. This indicated a structural relationship between subunits 4 and 6 during biogenesis of Fo. It therefore appears that subunit 4 (also called subunit b in beef heart and Escherichia coli ATP synthases) plays at least a structural role in the assembly of the whole complex. Disruption of the ATP4 gene also had a dramatic effect on the assembly of other mitochondrial complexes. Thus, the cytochrome oxidase activity of the mutant strain was about five times lower than that of the wild type. In addition, a high percentage of spontaneous rho-mutants was detected.ATP synthase is composed of subunits encoded by two genetic systems. In Saccharom,vces cerevisiae, the soluble portion of the enzyme, F1, contains the catalytic site for synthesis and hydrolysis of ATP. It is composed of five subunits (a, p, y, 6, E ) [l] that are encoded by nuclear DNA translated in the cytoplasm [2] and imported into the organelles [3]. The membranous portion of the complex, named Fo, catalyses H + conduction across the membrane and confers oligomycinsensitivity to F1 [4]. This membranous domain contains three highly hydrophobic mitochondrial gene products (subunits 6, 8 and 9) [5-81. Three additional nuclear-encoded polypeptides participate in the complex [l, 9-111 and presumably constitute a link between F1 and the three highly hydrophobic polypeptides of Fo. Electrophoresis shows their apparent molecular masses to be 25, 21 and 19 kDa respectively. In yeast, the structure of subunit 4 (25 kDa) [12] and the oligomycinsensitivity-conferring protein (21 kDa) [13] were recently reported.In previous papers, we have reported the purification procedure for subunit 4 and the sequence of the nuclear gene ATP4 [12, 141. Mature subunit 4 contains 209 amino acid residues/molecule and the predicted molecular mass is 23250 Da. A high similarity between subunit 4 and the b subunits of E. cofi and beef heart mitochondrial ATP synthases was reported [15].As revealed by secondary structure prediction methods, two distinct domains should compose subunit 4. The hydrophobic N-terminal part and the hydrophilic C-terminal sector Enzyme. ATP synthase or ATP phosphohydrolase (EC 3.6.1.34).confer amphiphilic behaviour to this subunit. Cross-linking experiments were previously reported between subunit 4 and F1 subunits c1 and b, and Fo subunits 9 and 6 [16, 171. Proteolytic cleavage of bovine submitochondrial particles depleted in F...

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“…It has been proposed that the F 1 sector biogenesis occurs independently of the F 0 sector (9) and requires the presence of the general Hsp60p chaperone and at least three assembly factors (Atp12p, Atp11p, and Fmc1p) with specific actions in the hexamerization of the ␣-F 1 and ␤-F 1 proteins (10 -12). Subunit 9 pre-formed oligomer in the inner mitochondrial membrane is believed to anchor the F 1 sector to the membrane (13), forming an F 1 -9 10 complex to which subunits 8, 4, and other F 0 components are assembled (14,15).…”

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confidence: 99%

Yeast Cells Depleted in Atp14p Fail to Assemble Atp6p within the ATP Synthase and Exhibit Altered Mitochondrial Cristae Morphology

Goyon¹,

Fronzes²,

Salin

et al. 2008

Journal of Biological Chemistry

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Within the yeast mitochondrial ATP synthase, subunit h is a small nuclear encoded protein belonging to the so-called "peripheral stalk" that connects the enzyme catalytic F 1 component to the mitochondrial inner membrane. This study examines the role of subunit h in ATP synthase function and assembly using a regulatable, doxycycline-repressible subunit h gene to overcome the strong instability of the mtDNA previously observed in strains lacking the native subunit h gene. Yeast cells expressing less than 3% of subunit h, but still containing intact mitochondrial genomes, grew poorly on respiratory substrates because of a major impairment of ATP synthesis originating from the ATP synthase, whereas the respiratory chain complexes were not affected. The lack of ATP synthesis in the subunit h-depleted (␦h) mitochondria was attributed to defects in the assembly/stability of the ATP synthase. A main feature of ␦h-mitochondria was a very low content (<6%) in the mitochondrially encoded Atp6p subunit, an essential component of the enzyme proton channel, which was in large part because of a slowing down in translation. Interestingly, depletion of subunit h resulted in dramatic changes in mitochondrial cristae morphology, which further supports the existence of a link between the ATP synthase and the folding/biogenesis of the inner mitochondrial membrane.

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Biogenesis of mitochondria: Defective yeast H+-ATPase assembled in the absence of mitochondrial protein synthesis is membrane associated

Cited by 16 publications

References 39 publications

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

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

The role of subunit 4, a nuclear‐encoded protein of the F₀ sector of yeast mitochondrial ATP synthase, in the assembly of the whole complex

Yeast Cells Depleted in Atp14p Fail to Assemble Atp6p within the ATP Synthase and Exhibit Altered Mitochondrial Cristae Morphology

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Biogenesis of mitochondria: Defective yeast H+-ATPase assembled in the absence of mitochondrial protein synthesis is membrane associated

Cited by 16 publications

References 39 publications

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

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

The role of subunit 4, a nuclear‐encoded protein of the F0 sector of yeast mitochondrial ATP synthase, in the assembly of the whole complex

Yeast Cells Depleted in Atp14p Fail to Assemble Atp6p within the ATP Synthase and Exhibit Altered Mitochondrial Cristae Morphology

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The role of subunit 4, a nuclear‐encoded protein of the F₀ sector of yeast mitochondrial ATP synthase, in the assembly of the whole complex