Phenotypic spectrum and clinical course of single large-scale mitochondrial DNA deletion disease in the paediatric population : a multicentre study

Background Large-scale mitochondrial DNA deletions (LMD) are a common genetic cause of mitochondrial disease and give rise to a wide range of clinical features. Lack of longitudinal data means the natural history remains unclear. This study was undertaken to describe the clinical spectrum in a large cohort of patients with paediatric disease onset. Methods A retrospective multicentre study was performed in patients with clinical onsetPeer reviewe

Crisis in Energy Metabolism - Mitochondrial Defects and a New Disease Entity

Impairment of energy metabolism may be associated with severe implications for affected individuals since all fundamental cell functions are energy-dependent. Disorders of energy metabolism are often genetic and associated with defects in the oxidative phosphorylation in mitochondria. This thesis addresses the pathogenesis in some mitochondrial disorders and a new disease entity associated with defects in the glycogen metabolism. In paper I we report on a primary mutation in mitochondrial DNA. We identified a T→C mutation at position 582 in the gene for tRNAPhe in a case of mitochondrial myopathy. The mutation alters a conserved base pairing in the aminoacyl stem of the tRNA. By analysis of single muscle fibers we showed that the level of heteroplasmy (proportion of mutant mtDNA) was higher in muscle fibers with defective cytochrome c oxidase (COX) activity compared to normal muscle fibers. Based on these findings we conclude that this mutation was responsible for the disease. In paper II we investigated a 30-year-old woman, who presented with an attack of acute rhabdomyolysis. We found an isolated deficiency of COX and a novel nonsense mutation in mtDNA in the gene encoding COX subunit I. In addition to its catalytic function, our data clearly indicates an important function of subunit I for the assembly of COX. The mutation was restricted to the patient’s muscle, but was not detectable in myoblasts, cultured from satellite cells isolated from affected muscle tissue. This result may have interesting implications for the natural evolution of the disease and perhaps therapy, since regenerating muscle occurs by proliferation of satellite cells. In paper III we investigated patients with mitochondrial diseases (progressive external ophthalmoplegia, PEO) due to primary mutations in POLG1 encoding mtDNA polymerase gamma (Polγ) and secondary multiple mtDNA deletions. The results show that it is very unlikely that mtDNA point mutations contribute to the pathogenesis in PEO patients with primary POLG1 mutations, and that the mechanism by which mutant Polγ cause mtDNA deletions does not involve mtDNA point mutations as an intermediate step, as has been previously proposed. In paper IV mtDNA alterations and pathology of muscle, brain and liver was investigated in children with Alpers-Huttenlocher syndrome (AHS), a fatal neurodegenerative disease associated with liver failure. All children had compound heterozygous missense mutations in POLG1. We provide evidence that AHS is a mitochondrial disease by demonstrating mtDNA alterations (reduced mtDNA copy number and multiple mtDNA deletions). Liver disease was triggered by valproate treatment in several cases possibly due to severe respiratory chain deficiency, which was demonstrated in liver tissue in one case. In paper V we report on a new disease entity “Muscle glycogen storage disease type zero” due to a homozygous stop mutation in the muscle glycogen synthase gene (GYS1). We performed investigations on a family where one child suffered sudden cardiac death at the age of 10 and his younger brother showed muscle fatigability and hypertrophic cardiomyopathy. In muscle there was a profound glycogen deficiency and an almost total predominance of oxidative muscle fibers

Identification of a large intronic transposal insertion in SLC17A5 causing sialic acid storage disease

Background: Sialic acid storage diseases are neurodegenerative disorders characterized by accumulation of sialic acid in the lysosome. These disorders are caused by mutations in SLC17A5, the gene encoding sialin, a sialic acid transporter located in the lysosomal membrane. The most common form of sialic acid storage disease is the slowly progressive Salla disease, presenting with hypotonia, ataxia, epilepsy, nystagmus and findings of cerebral and cerebellar atrophy. Hypomyelination and corpus callosum hypoplasia are typical as well. We report a 16 year-old boy with an atypically mild clinical phenotype of sialic acid storage disease characterized by psychomotor retardation and a mixture of spasticity and rigidity but no ataxia, and only weak features of hypomyelination and thinning of corpus callosum on MRI of the brain. Results: The thiobarbituric acid method showed elevated levels of free sialic acid in urine and fibroblasts, indicating sialic acid storage disease. Initial Sanger sequencing of SLC17A5 coding regions did not show any pathogenic variants, although exon 9 could not be sequenced. Whole exome sequencing followed by RNA and genomic DNA analysis identified a homozygous 6040 bp insertion in intron 9 of SLC17A5 corresponding to a long interspersed element-1 retrotransposon (KF425758.1). This insertion adds two splice sites, both resulting in a frameshift which in turn creates a premature stop codon 4 bp into intron 9. Conclusions: This study describes a novel pathogenic variant in SLC17A5, namely an intronic transposal insertion, in a patient with mild biochemical and clinical phenotypes. The presence of a small fraction of normal transcript may explain the mild phenotype. This case illustrates the importance of including lysosomal sialic acid storage disease in the differential diagnosis of developmental delay with postnatal onset and hypomyelination, as well as intronic regions in the genetic investigation of inborn errors of metabolism.Medicine, Faculty ofPharmaceutical Sciences, Faculty ofOther UBCNon UBCMedical Genetics, Department ofPediatrics, Department ofReviewedFacult

TANGO2 deficiency as a cause of neurodevelopmental delay with indirect effects on mitochondrial energy metabolism

Exome sequencing has recently identified mutations in the gene TANGO2 (transport and Golgi organization 2) as a cause of developmental delay associated with recurrent crises involving rhabdomyolysis, cardiac arrhythmias, and metabolic derangements. The disease is not well understood, in part as the cellular function and subcellular localization of the TANGO2 protein remain unknown. Furthermore, the clinical syndrome with its heterogeneity of symptoms, signs, and laboratory findings is still being defined. Here, we describe 11 new cases of TANGO2-related disease, confirming and further expanding the previously described clinical phenotype. Patients were homozygous or compound heterozygous for previously described exonic deletions or new frameshift, splice site, and missense mutations. All patients showed developmental delay with ataxia, dysarthria, intellectual disability, or signs of spastic diplegia. Of importance, we identify two subjects (aged 12 and 17 years) who have never experienced any overt episode of the catabolism-induced metabolic crises typical for the disease. Mitochondrial complex II activity was mildly reduced in patients investigated in association with crises but normal in other patients. In one deceased patient, post-mortem autopsy revealed heterotopic neurons in the cerebral white matter, indicating a possible role for TANGO2 in neuronal migration. Furthermore, we have addressed the subcellular localization of several alternative isoforms of TANGO2, none of which were mitochondrial but instead appeared to have a primarily cytoplasmic localization. Previously described aberrations in Golgi morphology were not observed in cultured skin fibroblasts

Additional file 1: of Identification of a large intronic transposal insertion in SLC17A5 causing sialic acid storage disease

Predicted sequences resulting from the LINE-1 retrotransposon insertion. (PDF 162 kb

Deoxyribonucleotide Metabolism in Cycling and Resting Human Fibroblasts with a Missense Mutation in p53R2, a Subunit of Ribonucleotide Reductase*

Ribonucleotide reduction provides deoxynucleotides for nuclear and mitochondrial (mt) DNA replication and DNA repair. In cycling mammalian cells the reaction is catalyzed by two proteins, R1 and R2. A third protein, p53R2, with the same function as R2, occurs in minute amounts. In quiescent cells, p53R2 replaces the absent R2. In humans, genetic inactivation of p53R2 causes early death with mtDNA depletion, especially in muscle. We found that cycling fibroblasts from a patient with a lethal mutation in p53R2 contained a normal amount of mtDNA and showed normal growth, ribonucleotide reduction, and deoxynucleoside triphosphate (dNTP) pools. However, when made quiescent by prolonged serum starvation the mutant cells strongly down-regulated ribonucleotide reduction, decreased their dCTP and dGTP pools, and virtually abolished the catabolism of dCTP in substrate cycles. mtDNA was not affected. Also, nuclear DNA synthesis and the cell cycle-regulated enzymes R2 and thymidine kinase 1 decreased strongly, but the mutant cell populations retained unexpectedly larger amounts of the two enzymes than the controls. This difference was probably due to their slightly larger fraction of S phase cells and therefore not induced by the absence of p53R2 activity. We conclude that loss of p53R2 affects ribonucleotide reduction only in resting cells and leads to a decrease of dNTP catabolism by substrate cycles that counterweigh the loss of anabolic activity. We speculate that this compensatory mechanism suffices to maintain mtDNA in fibroblasts but not in muscle cells with a larger content of mtDNA necessary for their high energy requirements

Molecular basis of infantile reversible cytochrome c oxidase deficiency myopathy.

Childhood-onset mitochondrial encephalomyopathies are usually severe, relentlessly progressive conditions that have a fatal outcome. However, a puzzling infantile disorder, long known as 'benign cytochrome c oxidase deficiency myopathy' is an exception because it shows spontaneous recovery if infants survive the first months of life. Current investigations cannot distinguish those with a good prognosis from those with terminal disease, making it very difficult to decide when to continue intensive supportive care. Here we define the principal molecular basis of the disorder by identifying a maternally inherited, homoplasmic m.14674T>C mt-tRNA(Glu) mutation in 17 patients from 12 families. Our results provide functional evidence for the pathogenicity of the mutation and show that tissue-specific mechanisms downstream of tRNA(Glu) may explain the spontaneous recovery. This study provides the rationale for a simple genetic test to identify infants with mitochondrial myopathy and good prognosis