Dataset Information

Mutation in subdomain G' of mitochondrial elongation factor G1 is associated with combined OXPHOS deficiency in fibroblasts but not in muscle.

ABSTRACT: The mitochondrial translation system is responsible for the synthesis of 13 proteins required for oxidative phosphorylation (OXPHOS), the major energy-generating process of our cells. Mitochondrial translation is controlled by various nuclear encoded proteins. In 27 patients with combined OXPHOS deficiencies, in whom complex II (the only complex that is entirely encoded by the nuclear DNA) showed normal activities, and mutations in the mitochondrial genome as well as polymerase gamma were excluded, we screened all mitochondrial translation factors for mutations. Here, we report a mutation in mitochondrial elongation factor G1 (GFM1) in a patient affected by severe, rapidly progressive mitochondrial encephalopathy. This mutation is predicted to result in an Arg250Trp substitution in subdomain G' of the elongation factor G1 protein and is presumed to hamper ribosome-dependent GTP hydrolysis. Strikingly, the decrease in enzyme activities of complex I, III and IV detected in patient fibroblasts was not found in muscle tissue. The OXPHOS system defects and the impairment in mitochondrial translation in fibroblasts were rescued by overexpressing wild-type GFM1, establishing the GFM1 defect as the cause of the fatal mitochondrial disease. Furthermore, this study evinces the importance of a thorough diagnostic biochemical analysis of both muscle tissue and fibroblasts in patients suspected to suffer from a mitochondrial disorder, as enzyme deficiencies can be selectively expressed.

SUBMITTER: Smits P

PROVIDER: S-EPMC3062000 | biostudies-literature | 2011 Mar

REPOSITORIES: biostudies-literature

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Mutation in subdomain G' of mitochondrial elongation factor G1 is associated with combined OXPHOS deficiency in fibroblasts but not in muscle.

Smits Paulien P Antonicka Hana H van Hasselt Peter M PM Weraarpachai Woranontee W Haller Wolfram W Schreurs Marieke M Venselaar Hanka H Rodenburg Richard J RJ Smeitink Jan A JA van den Heuvel Lambert P LP

European journal of human genetics : EJHG 20101201 3

The mitochondrial translation system is responsible for the synthesis of 13 proteins required for oxidative phosphorylation (OXPHOS), the major energy-generating process of our cells. Mitochondrial translation is controlled by various nuclear encoded proteins. In 27 patients with combined OXPHOS deficiencies, in whom complex II (the only complex that is entirely encoded by the nuclear DNA) showed normal activities, and mutations in the mitochondrial genome as well as polymerase gamma were exclud ...[more]

PMID: 21119709

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Project description:BackgroundCaveolin-3 (Cav3) is the principal structural component of caveolae in skeletal muscle. Dominant pathogenic mutations in the Cav3 gene, such as the Limb Girdle Muscular Dystrophy-1C (LGMD1C) P104L mutation, result in substantial loss of Cav3 and myopathic changes characterized by muscle weakness and wasting. We hypothesize such myopathy may also be associated with disturbances in mitochondrial biology. Herein, we report studies assessing the effects of Cav3 deficiency on mitochondrial form and function in skeletal muscle cells.MethodsL6 myoblasts were stably transfected with Cav3P104L or expression of native Cav3 repressed by shRNA or CRISPR/Cas9 genome editing prior to performing fixed/live cell imaging of mitochondrial morphology, subcellular fractionation and immunoblotting, or analysis of real time mitochondrial respiration. Skeletal muscle from wild-type and Cav3-/- mice was processed for analysis of mitochondrial proteins by immunoblotting.ResultsCaveolin-3 was detected in mitochondrial-enriched membranes isolated from mouse gastrocnemius muscle and L6 myoblasts. Expression of Cav3P104L in L6 myoblasts led to its targeting to the Golgi and loss of native Cav3 (>95%), including that associated with mitochondrial membranes. Cav3P104L reduced mitochondrial mass and induced fragmentation of the mitochondrial network that was associated with significant loss of proteins involved in mitochondrial biogenesis, respiration, morphology, and redox function [i.e. PGC1α, succinate dehyrdogenase (SDHA), ANT1, MFN2, OPA1, and MnSOD). Furthermore, Cav3P104L myoblasts exhibited increased mitochondrial cholesterol and loss of cardiolipin. Consistent with these changes, Cav3P104L expression reduced mitochondrial respiratory capacity and increased myocellular superoxide production. These morphological, biochemical, and functional mitochondrial changes were phenocopied in myoblasts in which Cav3 had been silenced/knocked-out using shRNA or CRISPR. Reduced mitochondrial mass, PGC1α, SDHA, ANT1, and MnSOD were also demonstrable in Cav3-/- mouse gastrocnemius. Strikingly, Cav3 re-expression in Cav3KO myoblasts restored its mitochondrial association and facilitated reformation of a tubular mitochondrial network. Significantly, re-expression also mitigated changes in mitochondrial superoxide, cholesterol, and cardiolipin content and recovered cellular respiratory capacity.ConclusionsOur results identify Cav3 as an important regulator of mitochondrial homeostasis and reveal that Cav3 deficiency in muscle cells associated with the Cav3P104L mutation invokes major disturbances in mitochondrial respiration and energy status that may contribute to the pathology of LGMD1C.

Dataset Information

Mutation in subdomain G' of mitochondrial elongation factor G1 is associated with combined OXPHOS deficiency in fibroblasts but not in muscle.

Publications

Mutation in subdomain G' of mitochondrial elongation factor G1 is associated with combined OXPHOS deficiency in fibroblasts but not in muscle.

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