Coi1 is a novel assembly factor of the yeast complex III-complex IV supercomplex.

The yeast bc1 complex (complex III) and cytochrome oxidase (complex IV) are mosaics of core subunits encoded by the mitochondrial genome and additional nuclear-encoded proteins imported from the cytosol. Both complexes build in the mitochondrial inner membrane various supramolecular assemblies. The formation of the individual complexes and their supercomplexes depends on the activity of dedicated assembly factors.

Mutational Analysis of the QRRQ Motif in the Yeast Hig1 Type 2 Protein Rcf1 Reveals a Regulatory Role for the Cytochrome Oxidase Complex.

The yeast Rcf1 protein is a member of the conserved family of proteins termed the hypoxia-induced gene (domain) 1 (Hig1 or HIGD1) family. Rcf1 interacts with components of the mitochondrial oxidative phosphorylation system, in particular the cytochrome (complex III)-cytochrome oxidase (complex IV) supercomplex (termed III-IV) and the ADP/ATP carrier proteins. Rcf1 plays a role in the assembly and modulation of the activity of complex IV; however, the molecular basis for how Rcf1 influences the activity of complex IV is currently unknown.

Cytochrome P450 enzymes but not NADPH oxidases are the source of the NADPH-dependent lucigenin chemiluminescence in membrane assays.

Unlabelled: Measuring NADPH oxidase (Nox)-derived reactive oxygen species (ROS) in living tissues and cells is a constant challenge. All probes available display limitations regarding sensitivity, specificity or demand highly specialized detection techniques. In search for a presumably easy, versatile, sensitive and specific technique, numerous studies have used NADPH-stimulated assays in membrane fractions which have been suggested to reflect Nox activity.

Mic13 Is Essential for Formation of Crista Junctions in Mammalian Cells.

Mitochondrial cristae are connected to the inner boundary membrane via crista junctions which are implicated in the regulation of oxidative phosphorylation, apoptosis, and import of lipids and proteins. The MICOS complex determines formation of crista junctions. We performed complexome profiling and identified Mic13, also termed Qil1, as a subunit of the MICOS complex.

Mutant desmin substantially perturbs mitochondrial morphology, function and maintenance in skeletal muscle tissue.

Secondary mitochondrial dysfunction is a feature in a wide variety of human protein aggregate diseases caused by mutations in different proteins, both in the central nervous system and in striated muscle. The functional relationship between the expression of a mutated protein and mitochondrial dysfunction is largely unknown. In particular, the mechanism how this dysfunction drives the disease process is still elusive.

Biallelic Mutations in TMEM126B Cause Severe Complex I Deficiency with a Variable Clinical Phenotype.

Complex I deficiency is the most common biochemical phenotype observed in individuals with mitochondrial disease. With 44 structural subunits and over 10 assembly factors, it is unsurprising that complex I deficiency is associated with clinical and genetic heterogeneity. Massively parallel sequencing (MPS) technologies including custom, targeted gene panels or unbiased whole-exome sequencing (WES) are hugely powerful in identifying the underlying genetic defect in a clinical diagnostic setting, yet many individuals remain without a genetic diagnosis.

Supercomplex-associated Cox26 protein binds to cytochrome c oxidase.

Here we identified a hydrophobic 6.4kDa protein, Cox26, as a novel component of yeast mitochondrial supercomplex comprising respiratory complexes III and IV. Multi-dimensional native and denaturing electrophoretic techniques were used to identify proteins interacting with Cox26.

MTO1-deficient mouse model mirrors the human phenotype showing complex I defect and cardiomyopathy.

Recently, mutations in the mitochondrial translation optimization factor 1 gene (MTO1) were identified as causative in children with hypertrophic cardiomyopathy, lactic acidosis and respiratory chain defect. Here, we describe an MTO1-deficient mouse model generated by gene trap mutagenesis that mirrors the human phenotype remarkably well. As in patients, the most prominent signs and symptoms were cardiovascular and included bradycardia and cardiomyopathy.

Cellular responses to egg-oil (charismon©).

Egg-oil (Charismon©) is known for its beneficial action in wound healing and other skin irritancies and its antibacterial activity. The physiological basis for these actions has been investigated using cells in culture: HaCaT-cells (immortalized human keratinocytes), human endothelial cells in culture (HUVEC), peripheral blood mononuclear lymphocytes (PBML) and a full thickness human skin model (FTSM). Emphasis was on the influence of egg-oil on cell migration and IL-8 production in HaCaT cells, respiration, mitochondrial membrane potential, reactive oxygen (ROS) production and proliferation in HUVEC and HaCaT cells, cytokine and interleukin production in PBML and UV-light induced damage of FTSM.

Phenotypic spectrum of eleven patients and five novel MTFMT mutations identified by exome sequencing and candidate gene screening.

Defects of mitochondrial oxidative phosphorylation (OXPHOS) are associated with a wide range of clinical phenotypes and time courses. Combined OXPHOS deficiencies are mainly caused by mutations of nuclear genes that are involved in mitochondrial protein translation. Due to their genetic heterogeneity it is almost impossible to diagnose OXPHOS patients on clinical grounds alone.

Quantification of protein complexes by blue native electrophoresis.

Blue native electrophoresis (BNE) is a long established method for the analysis of native protein complexes. Applications of BNE range from investigating subunit composition, stoichiometry, and assembly of single protein complexes to profiling of whole complexomes. BNE is an indispensible tool to diagnostically analyze cells and tissues from patients with mitochondrial disorders or model organisms.

Homozygous missense mutation in BOLA3 causes multiple mitochondrial dysfunctions syndrome in two siblings.

Defects of mitochondrial oxidative phosphorylation constitute a clinical and genetic heterogeneous group of disorders affecting multiple organ systems at varying age. Biochemical analysis of biopsy material demonstrates isolated or combined deficiency of mitochondrial respiratory chain enzyme complexes. Co-occurrence of impaired activity of the pyruvate dehydrogenase complex has been rarely reported so far and is not yet fully understood.

Molecular diagnosis in mitochondrial complex I deficiency using exome sequencing.

Background: Next generation sequencing has become the core technology for gene discovery in rare inherited disorders. However, the interpretation of the numerous sequence variants identified remains challenging. We assessed the application of exome sequencing for diagnostics in complex I deficiency, a disease with vast genetic heterogeneity.

Exome sequencing identifies ACAD9 mutations as a cause of complex I deficiency.

An isolated defect of respiratory chain complex I activity is a frequent biochemical abnormality in mitochondrial disorders. Despite intensive investigation in recent years, in most instances, the molecular basis underpinning complex I defects remains unknown. We report whole-exome sequencing of a single individual with severe, isolated complex I deficiency.

Respiratory chain complexes in dynamic mitochondria display a patchy distribution in life cells.

Background: Mitochondria, the main suppliers of cellular energy, are dynamic organelles that fuse and divide frequently. Constraining these processes impairs mitochondrial is closely linked to certain neurodegenerative diseases. It is proposed that functional mitochondrial dynamics allows the exchange of compounds thereby providing a rescue mechanism.

Large pore gels to separate mega protein complexes larger than 10 MDa by blue native electrophoresis: isolation of putative respiratory strings or patches.

Here, we expand the application of blue native electrophoresis to the separation of mega protein complexes larger than 10 MDa by introducing novel large pore acrylamide gels. We tailored the bis-acrylamide cross-linker amounts relative to the acrylamide monomer to enlarge the pore size of acrylamide gels and to obtain elastic and sufficiently stable gels. The novel gel types were then used to search for suprastructures of mitochondrial respiratory supercomplexes, the hypothetical respiratory strings, or patches.

Determination of protein mobility in mitochondrial membranes of living cells.

Molecular mobility in membranes of intracellular organelles is poorly understood, due to the lack of experimental tools applicable for a great diversity of shapes and sizes such organelles can acquire. Determinations of diffusion within the plasma membrane or cytosol are based mostly on the assumption of an infinite flat space, not valid for curved membranes of smaller organelles. Here we extend the application of FRAP to mitochondria of living cells by application of numerical analysis to data collected from a small region inside a single organelle.

Aging of different avian cultured cells: lack of ROS-induced damage and quality control mechanisms.

Elevated reactive oxygen species (ROS) levels have been observed in mammals during aging, implying an important role of ROS in the aging process. Most bird species are known to live longer and to contain lower ROS levels than mammals of the same body weight. The influence of ROS on the aging process of birds has been investigated using pigeon embryonic fibroblasts (PEF) and chicken embryonic fibroblasts (CEF).