The textbook description of mitochondrial respiratory complexes (RCs) views them as free-moving entities linked by the mobile carriers coenzyme Q (CoQ) and cytochrome c (cyt c). This model (known as the fluid model) is challenged by the proposal that all RCs except complex II can associate in supercomplexes (SCs). The proposed SCs are the respirasome (complexes I, III, and IV), complexes I and III, and complexes III and IV. The role of SCs is unclear, and their existence is debated. By genetic modulation of interactions between complexes I and III and III and IV, we show that these associations define dedicated CoQ and cyt c pools and that SC assembly is dynamic and organizes electron flux to optimize the use of available substrates.

The mitochondrial (mt) DNA depletion syndromes (MDDS) are genetic disorders characterized by a severe, tissue-specific decrease of mtDNA copy number, leading to organ failure. There are two main clinical presentations: myopathic (OMIM 609560) and hepatocerebral (OMIM 251880). Known mutant genes, including TK2, SUCLA2, DGUOK and POLG, account for only a fraction of MDDS cases. We found a new locus for hepatocerebral MDDS on chromosome 2p21-23 and prioritized the genes on this locus using a new integrative genomics strategy. One of the top-scoring candidates was the human ortholog of the mouse kidney disease gene Mpv17. We found disease-segregating mutations in three families with hepatocerebral MDDS and demonstrated that, contrary to the alleged peroxisomal localization of the MPV17 gene product, MPV17 is a mitochondrial inner membrane protein, and its absence or malfunction causes oxidative phosphorylation (OXPHOS) failure and mtDNA depletion, not only in affected individuals but also in Mpv17-/- mice.

Mitochondria are dynamic subcellular organelles that convert nutrient intermediates into readily available energy equivalents. Optimal mitochondrial function is ensured by a highly evolved quality control system, coordinated by protein machinery that regulates a process of continual fusion and fission. In this work, we provide in vivo evidence that the ATP-independent metalloprotease OMA1 plays an essential role in the proteolytic inactivation of the dynamin-related GTPase OPA1 (optic atrophy 1). We also show that OMA1 deficiency causes a profound perturbation of the mitochondrial fusion-fission equilibrium that has important implications for metabolic homeostasis. Thus, ablation of OMA1 in mice results in marked transcriptional changes in genes of lipid and glucose metabolic pathways and substantial alterations in circulating blood parameters. Additionally, Oma1-mutant mice exhibit an increase in body weight due to increased adipose mass, hepatic steatosis, decreased energy expenditure and impaired thermogenenesis. These alterations are especially significant under metabolic stress conditions, indicating that an intact OMA1-OPA1 system is essential for developing the appropriate adaptive response to different metabolic stressors such as a high-fat diet or cold-shock. This study provides the first description of an unexpected role in energy metabolism for the metalloprotease OMA1 and reinforces the importance of mitochondrial quality control for normal metabolic function.

The assembly of the five oxidative phosphorylation system (OXPHOS) complexes in the inner mitochondrial membrane is an intricate process. The human enzymes comprise core proteins, performing the catalytic activities, and a large number of ‘supernumerary’ subunits that play essential roles in assembly, regulation and stability. The correct addition of prosthetic groups as well as chaperoning and incorporation of the structural components require a large number of factors, many of which have been found mutated in cases of mitochondrial disease. Nowadays, the mechanisms of assembly for each of the individual complexes are almost completely understood and the knowledge about the assembly factors involved is constantly increasing. On the other hand, it is now well established that complexes I, III and IV interact with each other, forming the so-called respiratory supercomplexes or ‘respirasomes’, although the pathways that lead to their formation are still not completely clear. This review is a summary of our current knowledge concerning the assembly of complexes I–V and of the supercomplexes.

Mitochondrial protein translation is a complex process performed within mitochondria by an apparatus composed of mitochondrial DNA (mtDNA)-encoded RNAs and nuclear DNA-encoded proteins. Although the latter by far outnumber the former, the vast majority of mitochondrial translation defects in humans have been associated with mutations in RNA-encoding mtDNA genes, whereas mutations in protein-encoding nuclear genes have been identified in a handful of cases. Genetic investigation involving patients with defective mitochondrial translation led us to the discovery of novel mutations in the mitochondrial elongation factor G1 (EFG1) in one affected baby and, for the first time, in the mitochondrial elongation factor Tu (EFTu) in another one. Both patients were affected by severe lactic acidosis and rapidly progressive, fatal encephalopathy. The EFG1-mutant patient had early-onset Leigh syndrome, whereas the EFTu-mutant patient had severe infantile macrocystic leukodystrophy with micropolygyria. Structural modeling enabled us to make predictions about the effects of the mutations at the molecular level. Yeast and mammalian cell systems proved the pathogenic role of the mutant alleles by functional complementation in vivo. Nuclear-gene abnormalities causing mitochondrial translation defects represent a new, potentially broad field of mitochondrial medicine. Investigation of these defects is important to expand the molecular characterization of mitochondrial disorders and also may contribute to the elucidation of the complex control mechanisms, which regulate this fundamental pathway of mtDNA homeostasis.

Assembly of the oxidative phosphorylation (OXPHOS) system in the mitochondrial inner membrane is an intricate process in which many factors must interact. The OXPHOS system is composed of four respiratory chain complexes, which are responsible for electron transport and generation of the proton gradient in the mitochondrial intermembrane space, and of the ATP synthase that uses this proton gradient to produce ATP. Mitochondrial human disorders are caused by dysfunction of the OXPHOS system, and many of them are associated with altered assembly of one or more components of the OXPHOS system. The study of assembly defects in patients has been useful in unraveling and/or gaining a complete understanding of the processes by which these large multimeric complexes are formed. We review here current knowledge of the biogenesis of OXPHOS complexes based on investigation of the corresponding disorders.

Mitochondrial respiratory chain (MRC) enzymes associate in supercomplexes (SCs) that are structurally interdependent. This may explain why defects in a single component often produce combined enzyme deficiencies in patients. A case in point is the alleged destabilization of complex I in the absence of complex III. To clarify the structural and functional relationships between complexes, we have used comprehensive proteomic, functional, and biogenetical approaches to analyze a MT-CYB-deficient human cell line. We show that the absence of complex III blocks complex I biogenesis by preventing the incorporation of the NADH module rather than decreasing its stability. In addition, complex IV subunits appeared sequestered within complex III subassemblies, leading to defective complex IV assembly as well. Therefore, we propose that complex III is central for MRC maturation and SC formation. Our results challenge the notion that SC biogenesis requires the pre-formation of fully assembled individual complexes. In contrast, they support a cooperative-assembly model in which the main role of complex III in SCs is to provide a structural and functional platform for the completion of overall MRC biogenesis.

We investigated two unrelated children with an isolated defect of mitochondrial complex III activity. The clinical picture was characterized by a progressive encephalopathy featuring early-onset developmental delay, spasticity, seizures, lactic acidosis, brain atrophy and MRI signal changes in the basal ganglia. Both children were compound heterozygotes for novel mutations in the human bc1 synthesis like (BCS1L) gene, which encodes an AAA mitochondrial protein putatively involved in both iron homeostasis and complex III assembly. The pathogenic role of the mutations was confirmed by complementation assays, using a DeltaBcs1 strain of Saccharomyces cerevisiae. By investigating complex III assembly and the structural features of the BCS1L gene product in skeletal muscle, cultured fibroblasts and lymphoblastoid cell lines from our patients, we have demonstrated, for the first time in a mammalian system, that a major function of BCS1L is to promote the maturation of complex III and, more specifically, the incorporation of the Rieske iron-sulfur protein into the nascent complex. Defective BCS1L leads to the formation of a catalytically inactive, structurally unstable complex III. We have also shown that BCS1L is contained within a high-molecular-weight supramolecular complex which is clearly distinct from complex III intermediates.