Elsevier

Mitochondrion

Volume 18, September 2014, Pages 49-57
Mitochondrion

Riboflavin responsive mitochondrial myopathy is a new phenotype of dihydrolipoamide dehydrogenase deficiency. The chaperon-like effect of vitamin B2

https://doi.org/10.1016/j.mito.2014.09.006Get rights and content

Abstract

Dihydrolipoamide dehydrogenase (DLD, E3) is a flavoprotein common to pyruvate, α-ketoglutarate and branched-chain α-keto acid dehydrogenases. We found two novel DLD mutations (p.I40Lfs*4; p.G461E) in a 19 year-old patient with lactic acidosis and a complex amino- and organic aciduria consistent with DLD deficiency, manifesting progressive exertional fatigue. Muscle biopsy showed mitochondrial proliferation and lack of DLD cross-reacting material. Riboflavin supplementation determined the complete resolution of exercise intolerance with the partial restoration of the DLD protein and disappearance of mitochondrial proliferation in the muscle. Morphological and functional studies support the riboflavin chaperon-like role in stabilizing DLD protein with rescue of its expression in the muscle.

Introduction

Mammalian dihydrolipoamide dehydrogenase (dihydrolipoamide: NAD+ oxidoreductase, DLD, also designed as lipoamide dehydrogenase/LAD, EC 1.8.1.4) is the common flavoprotein component of three mitochondrial multi-enzyme complexes: pyruvate dehydrogenase complex (PDHc), α-ketoglutarate dehydrogenase complex (KGDHc) and branched-chain α-keto acid dehydrogenase complex (BCKDHc). DLD is also defined as E3, because within these complexes it catalyzes the third stage of each reaction (Cameron et al., 2006, Patel and Roche, 1990). DLD is also a component of the glycine cleavage system, another mitochondrial enzyme complex, where it is known as protein L (Kikuchi and Hiraga, 1982). In the context of these four multi-enzyme complexes, E3 utilizes dihydrolipoic acid and NAD+ to generate lipoic acid and NADH (Carothers et al., 1989). The active enzyme is a homodimer of 51 kDa subunits with four distinctive subdomain structures (FAD binding, NAD+ binding, central and interface domains) (Hong et al., 1997). The DLD gene, located on chromosome 7q31–32, is approximately 20 kb long and contains 14 exons (Grafakou et al., 2003).

Since the E3 is involved in different metabolic pathways within mitochondria, the clinical presentation of DLD deficiency (OMIM #246900) varies greatly, ranging from a severe neonatal presentation with hypoglycemia, ketoacidosis and encephalopathy or Leigh-like encephalopathy, psychomotor retardation and Reye-like syndrome to a milder presentation with exertional fatigue and myopathic features. The majority of patients become symptomatic within the first 1–2 years of age, and neonatal onset cases are associated with the worse prognosis; while some non-neonatal patients have been reported to survive into their second and third decades of age (Barak et al., 1998, Brassier et al., 2013, Quinonez et al., 2013, Quintana et al., 2010, Shaag et al., 1999). The wide clinical spectrum of DLD deficiency has also been attributed to the effects of different DLD mutations on the protein stability (Shaag et al., 1999) as well as on its ability to dimerize (Odièvre et al., 2005, Shany et al., 1999) or interact with other components of the three α-keto acid dehydrogenase complexes (Brautigam et al., 2006). E3 deficiency is often associated with an increased urinary excretion of α-keto acids, along with increased plasma levels of lactate and branched-chain amino acids leucine, valine and isoleucine.

We report on the clinical, morphological, biochemical and molecular findings of a 19-year-old young man presenting with muscle weakness and pain, intermittent elevation of blood lactate, ketoacidosis and creatine kinase. The patient is compound heterozygous for two novel pathogenic mutations in the DLD gene, and he was strikingly responsive to riboflavin treatment with the complete reversion of myopathic features.

Section snippets

Case report

This 19 year-old male patient was the first born of healthy Italian parents, following a spontaneous miscarriage in his mother. Neonatal parameters, weight, length and head circumference were in the normal range at birth. Early psychomotor development was uneventful, but hepatomegaly and recurrent episodes of drowsiness with ketonuria were noticed in infancy. At age 3 y he started with an inability to climb stairs together with episodes of muscle pain, weakness, and perioral cyanosis during

Molecular investigations

Sequence analysis of coding exon and flanking intronic sequences revealed that the patient was compound heterozygous for two novel mutations in DLD, the c.118 + 1G > T was inherited from the mother, and the c.1382G > A [p.G461E] inherited from the father (Fig. 1A). PCR amplification of cDNA with oligonucleotide primers encompassing the region of the c.118 + 1G > T mutation showed a lower band in association to the wild type (Fig. 1B); in fact, the mutation produces a skipping of exon 2 and predicts a

Discussion

DLD deficiency is a rare autosomal recessive metabolic disorder associated with variable phenotypic features. The metabolic profile of our patient was clearly consistent with E3 deficiency, showing the elevation of lactate and branched-chain amino acids in plasma (including alloisoleucine) with increased urinary excretion of α-keto acids. Differently from a recent paper reporting elevated plasma citrulline in DLD deficiency (Haviv et al., 2014), our patient had persistently normal levels of

Acknowledgments

This research was supported by grants from Telethon-Mitocon Project (Telethon GUP09004), and Ricerca Corrente (201302G002980) to EB.

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