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Screen for abnormal mitochondrial phenotypes in mouse embryonic stem cells identifies a model for succinyl-CoA ligase deficiency and mtDNA depletion.

ABSTRACT: Mutations in subunits of succinyl-CoA synthetase/ligase (SCS), a component of the citric acid cycle, are associated with mitochondrial encephalomyopathy, elevation of methylmalonic acid (MMA), and mitochondrial DNA (mtDNA) depletion. A FACS-based retroviral-mediated gene trap mutagenesis screen in mouse embryonic stem (ES) cells for abnormal mitochondrial phenotypes identified a gene trap allele of Sucla2 (Sucla2(SA?geo)), which was used to generate transgenic mice. Sucla2 encodes the ADP-specific ?-subunit isoform of SCS. Sucla2(SA?geo) homozygotes exhibited recessive lethality, with most mutants dying late in gestation (e18.5). Mutant placenta and embryonic (e17.5) brain, heart and muscle showed varying degrees of mtDNA depletion (20-60%). However, there was no mtDNA depletion in mutant liver, where the gene is not normally expressed. Elevated levels of MMA were observed in embryonic brain. SCS-deficient mouse embryonic fibroblasts (MEFs) demonstrated a 50% reduction in mtDNA content compared with wild-type MEFs. The mtDNA depletion resulted in reduced steady state levels of mtDNA encoded proteins and multiple respiratory chain deficiencies. mtDNA content could be restored by reintroduction of Sucla2. This mouse model of SCS deficiency and mtDNA depletion promises to provide insights into the pathogenesis of mitochondrial diseases with mtDNA depletion and into the biology of mtDNA maintenance. In addition, this report demonstrates the power of a genetic screen that combines gene trap mutagenesis and FACS analysis in mouse ES cells to identify mitochondrial phenotypes and to develop animal models of mitochondrial dysfunction.

SUBMITTER: Donti TR

PROVIDER: S-EPMC3917248 | biostudies-literature | 2014 Feb

REPOSITORIES: biostudies-literature

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Screen for abnormal mitochondrial phenotypes in mouse embryonic stem cells identifies a model for succinyl-CoA ligase deficiency and mtDNA depletion.

Donti Taraka R TR Stromberger Carmen C Ge Ming M Eldin Karen W KW Craigen William J WJ Graham Brett H BH

Disease models & mechanisms 20131121 2

Mutations in subunits of succinyl-CoA synthetase/ligase (SCS), a component of the citric acid cycle, are associated with mitochondrial encephalomyopathy, elevation of methylmalonic acid (MMA), and mitochondrial DNA (mtDNA) depletion. A FACS-based retroviral-mediated gene trap mutagenesis screen in mouse embryonic stem (ES) cells for abnormal mitochondrial phenotypes identified a gene trap allele of Sucla2 (Sucla2(SAβgeo)), which was used to generate transgenic mice. Sucla2 encodes the ADP-specif ...[more]

PMID: 24271779

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Project description:Mutations in SUCLA2 result in succinyl-CoA ligase (ATP-forming) or succinyl-CoA synthetase (ADP-forming) (A-SCS) deficiency, a mitochondrial tricarboxylic acid cycle disorder. The phenotype associated with this gene defect is largely encephalomyopathy. We describe two siblings compound heterozygous for SUCLA2 mutations, c.985A>G (p.M329V) and c.920C>T (p.A307V), with parents confirmed as carriers of each mutation. We developed a new LC-MS/MS based enzyme assay to demonstrate the decreased SCS activity in the siblings with this unique genotype. Both siblings shared bilateral progressive hearing loss, encephalopathy, global developmental delay, generalized myopathy, and dystonia with choreoathetosis. Prior to diagnosis and because of lactic acidosis and low activity of muscle pyruvate dehydrogenase complex (PDC), sibling 1 (S1) was placed on dichloroacetate, while sibling 2 (S2) was on a ketogenic diet. S1 developed severe cyclic vomiting refractory to therapy, while S2 developed Leigh syndrome, severe GI dysmotility, intermittent anemia, hypogammaglobulinemia and eventually succumbed to his disorder. The mitochondrial DNA contents in skeletal muscle (SM) were normal in both siblings. Pyruvate dehydrogenase complex, ketoglutarate dehydrogenase complex, and several mitochondrial electron transport chain (ETC) activities were low or at the low end of the reference range in frozen SM from S1 and/or S2. In contrast, activities of PDC, other mitochondrial enzymes of pyruvate metabolism, ETC and, integrated oxidative phosphorylation, in skin fibroblasts were not significantly impaired. Although we show that propionyl-CoA inhibits PDC, it does not appear to account for decreased PDC activity in SM. A better understanding of the mechanisms of phenotypic variability and the etiology for tissue-specific secondary deficiencies of mitochondrial enzymes of oxidative metabolism, and independently mitochondrial DNA depletion (common in other cases of A-SCS deficiency), is needed given the implications for control of lactic acidosis and possible clinical management.

Dataset Information

Screen for abnormal mitochondrial phenotypes in mouse embryonic stem cells identifies a model for succinyl-CoA ligase deficiency and mtDNA depletion.

Publications

Screen for abnormal mitochondrial phenotypes in mouse embryonic stem cells identifies a model for succinyl-CoA ligase deficiency and mtDNA depletion.

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