Metabolic disorders : genes and mechanisms

Sammanfattning: Mitochondrial disorders constitute a subgroup of the large family of rare monogenic diseases called Inborn Errors of Metabolism (IEM), but defects in mitochondrial activity have also been reported in more common and complex pathologies. In the present thesis, I validate new disease-causing variants and study the underlying molecular defects in three different cases of IEM. In paper I, I report the first individuals suffering from slowly-progressive, early-onset neurodegeneration caused by loss-of-function mutations in the autophagy adaptor Sequestosome 1 (SQSTM1, or p62). Analysis of fibroblasts from these patients suggests a defect in the early response to mitochondrial stress and in autophagosome formation. In paper II, I further investigate the phenotype of two of the previously reported patients, using reprogrammed neuroepithelial stem (NES) cells and differentiated neurones. Loss of p62 in NES cells leads to an impairment in neuronal differentiation, possibly due to an inability to switch between glycolytic and oxidative metabolism. In paper III, I study the consequences of the lack of Thioredoxin interacting protein (TXNIP) in myoblasts and fibroblasts of affected individuals. Oxygen consumption measurements revealed decreased mitochondrial activity, depending on the supplied carbon source. In particular, patient cells are unable to utilise the end product of glycolysis - pyruvate - but can efficiently use the tricarboxylic acid (TCA) cycle intermediate, malate, to fuel mitochondrial energy production. This expands the current view of the role of TXNIP in physiology and pathology. In paper IV, I model a neurodegenerative disorder related to mutations in the mitochondrial tryptophanyl-tRNA synthetase (WARS2) gene, after assessing pathogenicity of the variants in yeast. Knock down of WARS2 in Drosophila melanogaster resulted in larval lethality and reduced aminoacylation of tRNATrp, much resembling what can be observed in the affected individuals. Although muscle biopsies and fibroblasts from patients did not reveal a mitochondrial dysfunction, reprogrammed NES cells displayed a combined Complex I and IV defect, strengthening the relevant tissue-specificity of mitochondrial disorders. Our results demonstrate the importance of using appropriate models for the characterisation of mutations and for the efficient validation of pathogenic variants.