The inner membrane of the mitochondrion folds inwards, forming the cristae. This folding allows a greater amount of membrane to be packed into the mitochondrion. The data in this study demonstrate that subunits e and g of the mitochondrial ATP synthase are involved in generating mitochondrial cristae morphology. These two subunits are non-essential components of ATP synthase and are required for the dimerization and oligomerization of ATP synthase. Mitochondria of yeast cells de®cient in either subunits e or g were found to have numerous digitations and onion-like structures that correspond to an uncontrolled biogenesis and/or folding of the inner mitochondrial membrane. The present data show that there is a link between dimerization of the mitochondrial ATP synthase and cristae morphology. A model is proposed of the assembly of ATP synthase dimers, taking into account the oligomerization of the yeast enzyme and earlier data on the ultrastructure of mitochondrial cristae, which suggests that the association of ATP synthase dimers is involved in the control of the biogenesis of the inner mitochondrial membrane. Keywords: ATP synthase oligomer/mitochondria/ morphology/yeast IntroductionThe mitochondrion is referred to as the`power house' of the cell, because it is responsible for the synthesis of the majority of ATP under aerobic conditions. The inner membrane of the mitochondrion contains the components of the electron transport chain. Oxidation/reduction reactions along the components of the electron transport chain generate a proton gradient that is used by ATP synthase to phosphorylate ADP, thereby producing ATP. To increase the capacity of the mitochondrion to synthesize ATP, the inner membrane is folded to form cristae. These folds allow a much greater amount of electron transport chain enzymes and ATP synthase to be packed into the mitochondrion. However, until now, little was known about how the inner membrane is able to form cristae. This study provides evidence that subunits of ATP synthase are involved in cristae formation.ATP synthase, or F 1 F 0 ATP synthase, is composed of a hydrophilic catalytic unit (F 1 ), which is located in the mitochondrial matrix, and a membranous domain (F 0 ), which anchors the enzyme in the inner mitochondrial membrane and mediates the conduction of protons that participate indirectly in ATP synthesis (Fillingame, 1999;Pedersen et al., 2000). Electron microscopy of negatively stained mitochondria revealed 9 nm diameter projections in the mitochondrial matrix (Ferna Ândez-Mora Ân, 1962), which were identi®ed as the hydrophilic catalytic units (F 1 ) of the F 1 F 0 ATP synthase (Racker et al., 1965). These projections were observed by electron microscopy to be arranged in a non-random, tightly ordered pattern on tubular cristae in Paramecium multimicronucleatum mitochondria using rapid techniques of freezing followed by fracturing, etching and replication (Allen et al., 1989). In this organism, the F 1 complexes are arranged as a double row of particles along the full length ...

Atp6p is an essential subunit of the ATP synthase proton translocating domain, which is encoded by the mitochondrial DNA (mtDNA) in yeast. We have replaced the coding sequence of Atp6p gene with the non-respiratory genetic marker ARG8 m . Due to the presence of ARG8 m , accumulation of ؊ / 0 petites issued from large deletions in mtDNA could be restricted to 20 -30% by growing the atp6 mutant in media lacking arginine. This moderate mtDNA instability created favorable conditions to investigate the consequences of a specific lack in Atp6p. Interestingly, in addition to the expected loss of ATP synthase activity, the cytochrome c oxidase respiratory enzyme steadystate level was found to be extremely low (<5%) in the atp6 mutant. We show that the cytochrome c oxidase-poor accumulation was caused by a failure in the synthesis of one of its mtDNA-encoded subunits, Cox1p, indicating that, in yeast mitochondria, Cox1p synthesis is a key target for cytochrome c oxidase abundance regulation in relation to the ATP synthase activity. We provide direct evidence showing that in the absence of Atp6p the remaining subunits of the ATP synthase can still assemble. Mitochondrial cristae were detected in the atp6 mutant, showing that neither Atp6p nor the ATP synthase activity is critical for their formation. However, the atp6 mutant exhibited unusual mitochondrial structure and distribution anomalies, presumably caused by a strong delay in inner membrane fusion.In the mitochondrial inner membrane, the F 1 F 0 -type ATP synthase produces ATP from ADP and inorganic phosphate by using the energy of the transmembrane electrochemical proton gradient generated by the respiratory chain in the course of electron transfer to oxygen. The ATP synthase harbors two major structural domains, a transmembrane component (F 0 ) containing a proton-permeable pore and a peripheral, matrixlocalized, catalytic component (F 1 ) where the ATP is synthesized (1-4). In the F 0 , the core of the proton channel consists of a ring of c subunits (ten in yeast (4)) and one a subunit (Atp6p). Proton movement through this channel coincides with rotation of the subunit c ring (5-9), which results in conformational changes favoring ATP synthesis in the F 1 (1).Due to its good fermenting capacity the yeast Saccharomyces cerevisiae has been extensively used as a genetic system for the study of the mitochondrial ATP synthase (for reviews see Refs. 10 and 11). As in most eukaryotes, the yeast ATP synthase has a dual genetic origin, nuclear and mitochondrial. The yeast mitochondrial ATP synthase genes (ATP6, ATP9, and ATP8) encode the proton channel subunits a and c (usually referred to in yeast as Atp6p and Atp9p), respectively, and a third F 0 subunit (Atp8p) of unknown function. Dozens of mutations in the nuclear ATP synthase genes have provided much information on their protein products (10, 11). In contrast, only a very few mutants of the mitochondrial ATP synthase genes have been reported. Random generation of respiratory growth-deficient yeast strains issued fro...

We have identified a yeast nuclear gene (FMC1) that is required at elevated temperatures (37°C) for the formation/stability of the F 1 sector of the mitochondrial ATP synthase. Western blot analysis showed that Fmc1p is a soluble protein located in the mitochondrial matrix. At elevated temperatures in yeast cells lacking Fmc1p, the ␣-F 1 and ␤-F 1 proteins are synthesized, transported, and processed to their mature size. However, instead of being incorporated into a functional F 1 oligomer, they form large aggregates in the mitochondrial matrix. Identical perturbations were reported previously for yeast cells lacking either Atp12p or Atp11p, two specific assembly factors of the F 1 sector (Ackerman, S. H., and Tzagoloff, A. (1990) Proc. Natl. Acad. Sci. U. S. A. 87, 4986 -4990), and we show that the absence of Fmc1p can be efficiently compensated for by increasing the expression of Atp12p. However, unlike Atp12p and Atp11p, Fmc1p is not required in normal growth conditions (28 -30°C). We propose that Fmc1p is required for the proper folding/stability or functioning of Atp12p in heat stress conditions. F 1 F o -ATP synthases play a major role in cellular energy production. They are found in the plasma membranes of bacteria, thylakoid membranes of chloroplasts, and in the inner membrane of mitochondria. They use a proton gradient across their host membrane to produce ATP from ADP and inorganic phosphate (1, 2). This enzyme contains two distinct parts, called F o and F 1 . The F o mediates the transmembrane transport of protons, and the synthesis of ATP takes place on the F 1 .The F 1 contains five different types of subunits in the stoichiometric ratio ␣ 3 ␤ 3 ␥␦⑀ (3, 4). The three-dimensional structures of F 1 from bovine heart (5), rat liver (6) and yeast (7) show that the ␣-and ␤-subunits alternate in a hexagonal array with a central cavity occupied by the amino and carboxyl termini of the ␥-subunit. The interfaces between the ␣-and ␤-subunits form three catalytic and three noncatalytic nucleotide binding sites.In the yeast Saccharomyces cerevisiae, the F 1 subunits are encoded in the nucleus (8 -12), synthesized in the cytoplasm, imported into mitochondria as unfolded polypeptide chains (13), and then folded in the mitochondrial matrix with the help of Hsp60p and Hsp10p (14). The oligomerization of the F 1 monomers is assisted by two proteins called Atp12p and Atp11p. These interact directly with the ␣-F 1 and ␤-F 1 proteins, respectively (15, 16). In yeast strains lacking either Atp11p or Atp12p, the ␣-F 1 and ␤-F 1 proteins aggregate in the mitochondrial matrix (17). Thus it is believed that Atp12p and Atp11p facilitate the formation of ␣␤ heterodimers by protecting these two F 1 subunits from non-productive interactions (16).We report in this study the identification of Fmc1p, a novel protein required for the formation or stability of the F 1 oligomer. Like Atp11p and Atp12p, its absence also results in the aggregation of the ␣-F 1 and ␤-F 1 proteins. However, this is seen only at elevated temperatures (37°...

Mitochondrial diseases are severe and largely untreatable. Owing to the many essential processes carried out by mitochondria and the complex cellular systems that support these processes, these diseases are diverse, pleiotropic, and challenging to study. Much of our current understanding of mitochondrial function and dysfunction comes from studies in the baker's yeast Saccharomyces cerevisiae. Because of its good fermenting capacity, S. cerevisiae can survive mutations that inactivate oxidative phosphorylation, has the ability to tolerate the complete loss of mitochondrial DNA (a property referred to as ‘petite-positivity’), and is amenable to mitochondrial and nuclear genome manipulation. These attributes make it an excellent model system for studying and resolving the molecular basis of numerous mitochondrial diseases. Here, we review the invaluable insights this model organism has yielded about diseases caused by mitochondrial dysfunction, which ranges from primary defects in oxidative phosphorylation to metabolic disorders, as well as dysfunctions in maintaining the genome or in the dynamics of mitochondria. Owing to the high level of functional conservation between yeast and human mitochondrial genes, several yeast species have been instrumental in revealing the molecular mechanisms of pathogenic human mitochondrial gene mutations. Importantly, such insights have pointed to potential therapeutic targets, as have genetic and chemical screens using yeast.