Structure and Mechanism of Respiratory III-IV Supercomplexes in Bioenergetic Membranes.
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ABSTRACT:
SUBMITTER: Brzezinski P
PROVIDER: S-EPMC8361435 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description: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.
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Project description: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 analyse a MT-CYB-deficient human cell line. We show that 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.
| S-SCDT-EMBOJ-2019-102817 | biostudies-other
Project description:Mitochondria produce most of the ATP consumed by cells through the respiratory chain in their inner membrane. This process involves protein complexes assembled into larger structures, the respiratory supercomplexes (SCs). Cardiolipin (CL), the mitochondrial signature phospholipid, is crucial for the structural and functional integrity of these SCs, but it is as yet unclear by what mechanism it operates. Our data disclose the mechanism for bulk CL in gluing SCs, steering their formation, and suggest how it may stabilize specific interfaces. We describe self-assembly molecular dynamics simulations of 9 cytochrome bc1 (CIII) dimers and 27 cytochrome c oxidase (CIV) monomers from bovine heart mitochondria embedded in a CL-containing model lipid bilayer, aimed at mimicking the crowdedness and complexity of mitochondrial membranes. The simulations reveal a large diversity of interfaces, including those of existing experimental CIII/CIV SC models and an alternative interface with CIV rotated by 180°. SC interfaces enclose 4 to 12 CLs, a ?10 fold enrichment from the bulk. Half of these CLs glue complexes together using CL binding sites at the surface of both complexes. Free energy calculations demonstrate a larger CL binding strength, compared to other mitochondrial lipids, that is exclusive to these binding sites and results from non-additive electrostatic and van der Waals forces. This study provides a key example of the ability of lipids to selectively mediate protein-protein interactions by altering all ranges of forces, lubricate protein interfaces and act as traffic control agents steering proteins together.
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Project description:Mitochondrial respiratory chain complexes I, III, and IV can associate into larger structures termed supercomplexes or respirasomes, thereby generating structural interdependences among the individual complexes yet to be understood. In patients, nonsense mutations in complex IV subunit genes cause severe encephalomyopathies randomly associated with pleiotropic complex I defects. Using complexome profiling and biochemical analyses, we have explored the structural rearrangements of the respiratory chain in human cell lines depleted of the catalytic complex IV subunit COX1 or COX2. In the absence of a functional complex IV holoenzyme, several supercomplex I+III2 species coexist, which differ in their content of COX subunits and COX7A2L/HIGD2A assembly factors. The incorporation of an atypical COX1-HIGD2A submodule attenuates supercomplex I+III2 turnover rate, indicating an unexpected molecular adaptation for supercomplexes stabilization that relies on the presence of COX1 independently of holo-complex IV formation. Our data set the basis for complex I structural dependence on complex IV, revealing the co-existence of alternative pathways for the biogenesis of "supercomplex-associated" versus individual complex IV, which could determine physiological adaptations under different stress and disease scenarios.
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Project description:The organization of the mitochondrial electron transport chain proteins into supercomplexes (SCs) is now undisputed; however, their assembly process, or the role of differential expression isoforms, remain to be determined. In Saccharomyces cerevisiae, cytochrome c oxidase (CIV) forms SCs of varying stoichiometry with cytochrome bc 1 (CIII). Recent studies have revealed, in normoxic growth conditions, an interface made exclusively by Cox5A, the only yeast respiratory protein that exists as one of two isoforms depending on oxygen levels. Here we present the cryo-EM structures of the III2-IV1 and III2-IV2 SCs containing the hypoxic isoform Cox5B solved at 3.4 and 2.8 Å, respectively. We show that the change of isoform does not affect SC formation or activity, and that SC stoichiometry is dictated by the level of CIII/CIV biosynthesis. Comparison of the CIV5B- and CIV5A-containing SC structures highlighted few differences, found mainly in the region of Cox5. Additional density was revealed in all SCs, independent of the CIV isoform, in a pocket formed by Cox1, Cox3, Cox12, and Cox13, away from the CIII-CIV interface. In the CIV5B-containing hypoxic SCs, this could be confidently assigned to the hypoxia-induced gene 1 (Hig1) type 2 protein Rcf2. With conserved residues in mammalian Hig1 proteins and Cox3/Cox12/Cox13 orthologs, we propose that Hig1 type 2 proteins are stoichiometric subunits of CIV, at least when within a III-IV SC.
| S-EPMC7196821 | biostudies-literature
Project description:Mitochondrial respiratory chain complexes I, III and IV can associate into larger structures termed supercomplexes or respirasomes, thereby generating structural interdependences among the individual complexes yet to be understood. In patients, nonsense mutations in complex IV subunit genes cause severe encephalomyopathies randomly associated with pleiotropic complex I defects. Using complexome profiling and biochemical analyses, we have explored the structural rearrangements of the respiratory chain in human cell lines depleted of the catalytic complex IV subunits COX1 or COX2. In the absence of a functional complex IV holoenzyme, several supercomplex I+III2 species coexist, which differ in their content of COX subunits and COX7A2L/HIGD2A assembly factors. The incorporation of an atypical COX1-HIGD2A submodule attenuates supercomplex I+III2 turnover rate, indicating an unexpected molecular adaptation for supercomplexes stabilization that relies on the presence of COX1 independently of holo-complex IV formation. Our data set the basis for complex I structural dependence on complex IV, revealing the co-existence of alternative pathways for the biogenesis of 'supercomplex-associated' versus individual complex IV, which could determine physiological adaptations under different stress and disease scenarios.
| S-SCDT-EMBOJ-2019-103912 | biostudies-other
Project description:The role of OXPHOS respiratory supercomplexes remains still unknown. To answer this question we use as a model a SCAF1 null allele model in zebrafish. SCAF1 (cox7a2l) is the only supercomplex assembly factor identified in vertebrates and it is involved in the physical link of III-IV complexes. Therefore the SCAF1 null allele model lack all III-IV interactions. We demonstrate that supercomplexes provide an advantage in the optimization of metabolic resources.
2020-05-03 | GSE133487 | GEO
Project description:The respiratory complexes are organized in supramolecular assemblies called supercomplexes thought to optimize cellular metabolism under physiological and pathological conditions. In this study, we used genetically and biochemically well characterized cells bearing the pathogenic microdeletion m.15,649-15,666 (ΔI300-P305) in MT-CYB gene, to investigate the effects of an assembly-hampered CIII on the re-organization of supercomplexes. First, we found that this mutation also affects the stability of both CI and CIV, and evidences the occurrence of a preferential structural interaction between CI and CIII2, yielding a small amount of active CI+CIII2 supercomplex. Indeed, a residual CI+CIII combined redox activity, and a low but detectable ATP synthesis driven by CI substrates are detectable, suggesting that the assembly of CIII into the CI+CIII2 supercomplex mitigates the detrimental effects of MT-CYB deletion. Second, measurements of oxygen consumption and ATP synthesis driven by NADH-linked and FADH2-linked substrates alone, or in combination, indicate a common ubiquinone pool for the two respiratory pathways. Finally, we report that prolonged incubation with rotenone enhances the amount of CI and CIII2, but reduces CIV assembly. Conversely, the antioxidant N-acetylcysteine increases CIII2 and CIV2 and partially restores respirasome formation. Accordingly, after NAC treatment, the rate of ATP synthesis increases by two-fold compared with untreated cell, while the succinate level, which is enhanced by the homoplasmic mutation, markedly decreases. Overall, our findings show that fine-tuning the supercomplexes stability improves the energetic efficiency of cells with the MT-CYB microdeletion.
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Project description:The existence of respiratory supercomplexes (SCs) has been well acknowledged especially with the development of cryo-EM in the past few years, but their physiological relevance remains elusive. Dynamics of OXPHOS complexes is tightly correlated to cell fate switch, but whether SCs involve in this process is unknown. During C2C12 myogenesis, CIII2 and CIV as well as the subunits of CIII and CIV were slightly increasing, while SC III2+IV was decreasing, which indicates that SC reorganization may regulate myogenesis. Knockdown Cox7a2l decreased SCs III2+IV, I+III2+IV2 and I+III2+IV3, but had little effect on I+III2+IV. Transcriptome and quantitative-proteome analysis indicated that Cox7a2l silence activated myogenic gene expression and repressed cell cycle-associated gene expression. Besides, knockdown of Cox7a2l decreased CII abundance and accumulated succinate, which in turn suppressed certain key transcription factors, such as HMGA1 which is important for stemness maintenance. Our results indicated that respiratory supercomplex reorganization changed succinate homeostasis and myogenesis related gene expression profiles, thus switching cell fate.
2021-11-01 | GSE178547 | GEO
Project description:The organization of individual respiratory Complexes I, III, and IV (mammalian cells) or III and IV (yeast) of the mitochondria into higher order supercomplexes (SCs) is generally accepted. However, the factors that regulate SC formation and the functional significance of SCs are not well understood. The mitochondrial signature phospholipid cardiolipin (CL) plays a central role in formation and stability of respiratory SCs from yeast to man. Studies in yeast mutants in which the CL level can be regulated displayed a direct correlation between CL levels and SC formation. Disease states in which CL levels are reduced also show defects in SC formation. Three-dimensional density maps of yeast and bovine SCs by electron cryo-microscopy show gaps between the transmembrane-localized interfaces of individual complexes consistent with the large excess of CL in SCs over that integrated into the structure of individual respiratory complexes. Finally, the yeast SC composed of Complex III and two Complexes IV was reconstituted in liposomes from purified individual complexes containing integrated CLs. Reconstitution was wholly dependent on inclusion of additional CL in the liposomes. Therefore, non-integral CL molecules play an important role in SC formation and may be involved in regulation of SC stability under metabolic conditions where CL levels fluctuate.
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