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Expression of the alternative oxidase complements cytochrome c oxidase deficiency in human cells.

ABSTRACT: Cytochrome c oxidase (COX) deficiency is associated with a wide spectrum of clinical conditions, ranging from early onset devastating encephalomyopathy and cardiomyopathy, to neurological diseases in adulthood and in the elderly. No method of compensating successfully for COX deficiency has been reported so far. In vitro, COX-deficient human cells require additional glucose, pyruvate and uridine for normal growth and are specifically sensitive to oxidative stress. Here, we have tested whether the expression of a mitochondrially targeted, cyanide-resistant, alternative oxidase (AOX) from Ciona intestinalis could alleviate the metabolic abnormalities of COX-deficient human cells either from a patient harbouring a COX15 pathological mutation or rendered deficient by silencing the COX10 gene using shRNA. We demonstrate that the expression of the AOX, well-tolerated by the cells, compensates for both the growth defect and the pronounced oxidant-sensitivity of COX-deficient human cells.

SUBMITTER: Dassa EP

PROVIDER: S-EPMC3378104 | biostudies-literature | 2009 Apr

REPOSITORIES: biostudies-literature

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Expression of the alternative oxidase complements cytochrome c oxidase deficiency in human cells.

Dassa Emmanuel P EP Dufour Eric E Gonçalves Sérgio S Paupe Vincent V Hakkaart Gertjan A J GA Jacobs Howard T HT Rustin Pierre P

EMBO molecular medicine 20090401 1

Cytochrome c oxidase (COX) deficiency is associated with a wide spectrum of clinical conditions, ranging from early onset devastating encephalomyopathy and cardiomyopathy, to neurological diseases in adulthood and in the elderly. No method of compensating successfully for COX deficiency has been reported so far. In vitro, COX-deficient human cells require additional glucose, pyruvate and uridine for normal growth and are specifically sensitive to oxidative stress. Here, we have tested whether th ...[more]

PMID: 20049701

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Project description:BackgroundSirtuins are NAD+ dependent deacetylases, which regulate mitochondrial energy metabolism as well as cellular response to stress. The NAD/NADH-system plays a crucial role in oxidative phosphorylation linking sirtuins and the mitochondrial respiratory chain. Furthermore, sirtuins are able to directly deacetylate and activate different complexes of the respiratory chain. This prompted us to analyse sirtuin levels in skin fibroblasts from patients with cytochrome c-oxidase (COX) deficiency and to test the impact of different pharmaceutical activators of sirtuins (SRT1720, paeonol) to modulate sirtuins and possibly respiratory chain enzymes in patient cells in vitro.MethodsWe assayed intracellular levels of sirtuin 1 and the mitochondrial sirtuins SIRT3 and SIRT4 in human fibroblasts from patients with COX- deficiency. Furthermore, sirtuins were measured after inhibiting complex IV in healthy control fibroblasts by cyanide and after incubation with activators SRT1720 and paeonol. To determine the effect of sirtuin inhibition at the cellular level we measured total cellular acetylation (control and patient cells, with and without treatment) by Western blot.ResultsWe observed a significant decrease in cellular levels of all three sirtuins at the activity, protein and transcriptional level (by 15% to 50%) in COX-deficient cells. Additionally, the intracellular concentration of NAD+ was reduced in patient cells. We mimicked the biochemical phenotype of COX- deficiency by incubating healthy fibroblasts with cyanide and observed reduced sirtuin levels. A pharmacological activation of sirtuins resulted in normalized sirtuin levels in patient cells. Hyper acetylation was also reversible after treatment with sirtuin activators. Pharmacological modulation of sirtuins resulted in altered respiratory chain complex activities.ConclusionsWe found inhibition of situins 1, 3 and 4 at activity, protein and transcriptional levels in fibroblasts from patient with COX-deficiency. Pharmacological activators were able to restore reduced sirtuin levels and thereby modulate respiratory chain activities.

Dataset Information

Expression of the alternative oxidase complements cytochrome c oxidase deficiency in human cells.

Publications

Expression of the alternative oxidase complements cytochrome c oxidase deficiency in human cells.

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