Succinyl-CoA synthetase (SUCLA2) deficiency in two siblings with impaired activity of other mitochondrial oxidative enzymes in skeletal muscle without mitochondrial DNA depletion

Abstract

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.

Keywords: Electron transport chain complexes; Liquid chromatography tandem mass spectrometry; Propionyl-CoA; Pyruvate dehydrogenase complex deficiency; SUCLA2; Succinyl-CoA synthetase deficiency.

Copyright © 2016 Elsevier Inc. All rights reserved.

Figures

Fig. 1

Summary of the WES pipeline used in this work. MS, missense; FS, frameshift; SS, splice site; and X, stop.

Fig. 2

A-SCS activity measurement by LC-MS/MS. (A) Time dependency was measured in control line and patient line at 37°C for 0, 3, 6, 9 and 12 min with 50 mM Tris buffer pH 8.0, 5 mM MgCl2, 10 mM D4-succinate, 1 mM ATP, 1 mM CoA, oligomycin 2 µg/ml and homogenate protein concentration 1mg/ml. Activity linearity is shown within 9min. (B) Protein dependency was measured in control line for 6 min at different protein concentrations, 0, 0.5, 0.75, 1 and 1.25 mg/ml. Other conditions were same as (A). Linear for protein concentrations <1.25 mg/ml. (C) and (D) Apparent Km and Vmax of succinate were measured from two control lines with different D4-succinate concentrations, 0.37, 1.11, 3.33, 10 and 30 mM, and 1mg/ml homogenate incubated at 37°C for 6min. Other conditions were same as (A).

Fig. 3

Western blot analyses result of decreased SUCLA2 in patient lines. SUCLA2 protein detected by Western Blot and normalized to total GAPDH levels as loading controls. Relative SUCLA2 amounts were calculated using mean value of controls, indicated as percentages.

Fig. 4

Inhibition of PDC by propionyl-CoA. Vmax (nmol/min/mg protein) of PDC (A, purified porcine heart complex); or apparent Vmax (B, complex in disrupted cultured fibroblasts; and C, complex in SM homogenate) vs propionyl-CoA concentration (mM) and their respective determined Ki or apparent Ki