Mitochondrial cytochrome c oxidase deficiency

Abstract

As with other mitochondrial respiratory chain components, marked clinical and genetic heterogeneity is observed in patients with a cytochrome c oxidase deficiency. This constitutes a considerable diagnostic challenge and raises a number of puzzling questions. So far, pathological mutations have been reported in more than 30 genes, in both mitochondrial and nuclear DNA, affecting either structural subunits of the enzyme or proteins involved in its biogenesis. In this review, we discuss the possible causes of the discrepancy between the spectacular advances made in the identification of the molecular bases of cytochrome oxidase deficiency and the lack of any efficient treatment in diseases resulting from such deficiencies. This brings back many unsolved questions related to the frequent delay of clinical manifestation, variable course and severity, and tissue-involvement often associated with these diseases. In this context, we stress the importance of studying different models of these diseases, but also discuss the limitations encountered in most available disease models. In the future, with the possible exception of replacement therapy using genes, cells or organs, a better understanding of underlying mechanism(s) of these mitochondrial diseases is presumably required to develop efficient therapy.

Keywords: cytochrome c oxidase; genetic diseases; mitochondria; mitochondrial diseases; mtDNA; signalling pathway.

Figure 1. Cytochrome oxidase location in the respiratory chain and activity assay in human skin fibroblasts

A. Schematic representation of the respiratory chain in the inner mitochondrial membrane showing the interaction of cytochrome oxidase (complex IV) with complexes I and III in a super-complex (respirasome). The site of action of specific inhibitors is indicated in red. The green arrow shows the alternative oxidase (AOX) by-pass, which when expressed in COX-defective human mitochondria or flies rescues their various phenotypes. The assay of COX with externally added cytochrome c requires the permeabilization of the outer membrane. B. Cytochrome oxidase is assayed spectrophotometrically by measuring using a double-wavelength spectrophotometer (550–540 nm) the oxidation of reduced cytochrome c in skin fibroblasts permeabilized by 2 successive freeze/thaw cycles. The reaction is first order with respect to substrate concentration and is thus diminished by half when half of the reduced cytochrome c is consumed. Subsequent sequential addition of rotenone, cyanide, oxidized cytochrome c and succinate measures reduction of cytochrome c, first by the succinate-cytochrome c reductase (CII plus CIII). The activity is essentially rate controlled by CII and can be inhibited by malonate, a competitive inhibitor of CII. Further addition of glycerol-3 phosphate measures the activity from the glycerol-3 phosphate dehydrogenase (G3Pdh) to CIII. This activity can be selectively inhibited by iGP1(143). Finally, addition of decylubiquinol in the presence of EDTA is used to measure antimycin-sensitive CIII activity. Abbreviations: The RC complexes are abbreviated as, CI, CII, CIII, CIV, and the ATP synthase as CV; c, cytochrome c; COX, cytochrome oxidase; Ddh, the dihydroorotate dehydrogenase which catalyze the production of uridine, an essential step for the synthesis of nucleic acids; EDTA, ethylenediamine tetraacetic acid; ETF, the electron transfer flavoprotein involved in the oxidation of fatty acids; G3Pdh, the glycerol 3-phosophate dehydrogenase; GCCR, iGP1-sensitive glycerol 3-phosphate; IM, inner membrane; KCN, potassium cyanide; OM, outer membrane; QCCR, antimycin-sensitive decylubiquinol-cytochrome c reductase; SCCR, malonate-sensitive cytochrome c reductase; UQ, ubiquinone 50, or coenzyme Q₁₀.

Figure 2. Synthesis and assembly of COX subunits

A scheme summarizing what is presently known about the pathways for the integrated synthesis and assembly of COX subunits expressed from the nuclear and mitochondrial genomes. Protein subunits translated on cytosolic ribosomes with N-terminal presequences are first transported by the outer membrane TOM complex and are subsequently matured and sorted by the TIM and MITRAC machineries to the intermembrane space, inner membrane, or matrix where they interact with partner proteins to form assembly intermediates (–146). Subunits encoded by mtDNA genes are translated on mitochondrial ribosomes attached to the matrix side of the inner membrane. Following insertion into the inner membrane by Oxa1 they interact with their nucleo-cytoplasmic partners to form subcomplexes that subsequently assemble into COX. The overall process is assisted by numerous proteins acting in transport, translation, chaperoning of different assembly steps. Oxa1 is also involved in the biogenesis of other respiratory chain complexes. Some of the genes coding for ancillary factors (indicated in red) have been found to be mutated in COX deficient patients. IM, inner membrane; OM, outer membrane; 1, 2, 4, 5a, COX subunits (purple)

Figure 3. Maturation and insertion of COX into the respiratory chain

Mammalian COX exists as a dimer. Each monomer consists of 13 different subunits. At present human mutations leading to a COX deficiency have been identified in six structural subunits including the three mtDNA-encoded core proteins (in red) and in 9 ancillary proteins (also indicated in red). The catalytic activity of COX depends on heme a, a₃ and two copper centers (Cu_A and Cu_B) linking COX biosynthesis to both copper and heme metabolism. Maturation of COX active centers involves a number of factors, some of which have also been found mutated in COX deficient patients (denoted in red). IM, inner membrane; OM, outer membrane; 1, 2, 4, 5a, 5c, 6a, 6b, 6c, 7a, 7b, 8, the different COX subunits; CI, CII, CIII, CIV, the various complexes of the respiratory chain

Figure 4. The non-overlapping clinical symptoms of COX deficiency and expression territories of COX subunits

On the left are shown ubiquitously expressed subunits which when mutated result in a constellation of symptom but sparing numerous organs. The representations in the middle and on the right show that mutation in genes encoding subunits with more specific organ/tissue expression does not necessarily result in symptoms affecting the predicted organs. Subunits encoded in mtDNA are depicted in green. Genes with identified mutations in patients with COX deficiency are shown in yellow (or green).