Physiological diversity of mitochondrial oxidative phosphorylation.
ABSTRACT To investigate the physiological diversity in the regulation and control of mitochondrial oxidative phosphorylation, we determined the composition and functional features of the respiratory chain in muscle, heart, liver, kidney, and brain. First, we observed important variations in mitochondrial content and infrastructure via electron micrographs of the different tissue sections. Analyses of respiratory chain enzyme content by Western blot also showed large differences between tissues, in good correlation with the expression level of mitochondrial transcription factor A and the activity of citrate synthase. On the isolated mitochondria, we observed a conserved molar ratio between the respiratory chain complexes and a variable stoichiometry for coenzyme Q and cytochrome c, with typical values of [1-1.5]:[30-135]::[9-35]:[6.5-7.5] for complex II:coenzyme Q:complex III:cytochrome c:complex IV in the different tissues. The functional analysis revealed important differences in maximal velocities of respiratory chain complexes, with higher values in heart. However, calculation of the catalytic constants showed that brain contained the more active enzyme complexes. Hence, our study demonstrates that, in tissues, oxidative phosphorylation capacity is highly variable and diverse, as determined by different combinations of 1) the mitochondrial content, 2) the amount of respiratory chain complexes, and 3) their intrinsic activity. In all tissues, there was a large excess of enzyme capacity and intermediate substrate concentration, compared with what is required for state 3 respiration. To conclude, we submitted our data to a principal component analysis that revealed three groups of tissues: muscle and heart, brain, and liver and kidney.
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ABSTRACT: Hearts are the first organs to fail in animals exposed to heat stress. Predictions of climate change mediated increases in ocean temperatures suggest that the ectothermic heart may place tight constraints on the diversity and distribution of marine species with cardiovascular systems. For many such species, their upper temperature limits (Tmax) and respective heart failure (HF) temperature (THF) are only a few degrees from current environmental temperatures. While the ectothermic cardiovascular system acts as an "ecological thermometer," the exact mechanism that mediates HF remains unresolved. We propose that heat-stressed cardiac mitochondria drive HF. Using a common New Zealand fish, Notolabrus celidotus, we determined the THF (27.5°C). Haemoglobin oxygen saturation appeared to be unaltered in the blood surrounding and within heat stressed hearts. Using high resolution respirometry coupled to fluorimeters, we explored temperature-mediated changes in respiration, ROS and ATP production, and overlaid these changes with THF. Even at saturating oxygen levels several mitochondrial components were compromised before THF. Importantly, the capacity to efficiently produce ATP in the heart is limited at 25°C, and this is prior to the acute THF for N. celidotus. Membrane leakiness increased significantly at 25°C, as did cytochrome c release and permeability to NADH. Maximal flux rates and the capacity for the electron transport system to uncouple were also altered at 25°C. These data indicate that mitochondrial membrane integrity is lost, depressing ATP synthesis capacity and promoting cytochrome c release, prior to THF. Mitochondria can mediate HF in heat stressed hearts in fish and play a significant role in thermal stress tolerance, and perhaps limit species distributions by contributing to HF.PLoS ONE 01/2013; 8(5):e64120. · 3.73 Impact Factor
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ABSTRACT: Hereditary spastic paraplegia (HSP) is considered one of the most heterogeneous groups of neurological disorders, both clinically and genetically. The disease comprises pure and complex forms that clinically include slowly progressive lower-limb spasticity resulting from degeneration of the corticospinal tract. At least 48 loci accounting for these diseases have been mapped to date, and mutations have been identified in 22 genes, most of which play a role in intracellular trafficking. Here, we identified mutations in two functionally related genes (DDHD1 and CYP2U1) in individuals with autosomal-recessive forms of HSP by using either the classical positional cloning or a combination of whole-genome linkage mapping and next-generation sequencing. Interestingly, three subjects with CYP2U1 mutations presented with a thin corpus callosum, white-matter abnormalities, and/or calcification of the basal ganglia. These genes code for two enzymes involved in fatty-acid metabolism, and we have demonstrated in human cells that the HSP pathophysiology includes alteration of mitochondrial architecture and bioenergetics with increased oxidative stress. Our combined results focus attention on lipid metabolism as a critical HSP pathway with a deleterious impact on mitochondrial bioenergetic function.The American Journal of Human Genetics 11/2012; · 11.20 Impact Factor
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ABSTRACT: Mitochondrial respiratory chain is organised into supramolecular structures that can be preserved in mild detergent solubilisates and resolved by native electrophoretic systems. Supercomplexes of respiratory complexes I, III and IV as well as multimeric forms of ATP synthase are well established. However, the involvement of complex II, linking respiratory chain with tricarboxylic acid cycle, in mitochondrial supercomplexes is questionable. Here we show that digitonin-solubilised complex II quantitatively forms high molecular weight structures (CIIhmw) that can be resolved by clear native electrophoresis. CIIhmw structures are enzymatically active and differ in electrophoretic mobility between tissues (500 - over 1000 kDa) and cultured cells (400-670 kDa). While their formation is unaffected by isolated defects in other respiratory chain complexes, they are destabilised in mtDNA-depleted, rho0 cells. Molecular interactions responsible for the assembly of CIIhmw are rather weak with the complexes being more stable in tissues than in cultured cells. While electrophoretic studies and immunoprecipitation experiments of CIIhmw do not indicate specific interactions with the respiratory chain complexes I, III or IV or enzymes of the tricarboxylic acid cycle, they point out to a specific interaction between CII and ATP synthase.PLoS ONE 01/2013; 8(8):e71869. · 3.73 Impact Factor