Studies on the genetics and molecular pathogenesis of mitochondrial respiratory chain disorders

This work aimed to identify the molecular genetic basis of disease in thirty patients with classical mitochondrial encephalomyopathy phenotypes without known mutations. The patients exhibited a range of phenotypes including MELAS, MERRF, CPEO, exercise intolerance, isolated myopathy and Kearns-Sayre syndrome. In addition the molecular mechanisms which underlie the clinical diversity associated with the common A3243G and 7472C insertion- mutations were investigated. Systematic sequencing of the entire muscle mtDNA was undertaken in each of the thirty patients. Seven pathogenic mtDNA mutations were identified in eleven of the patients. Five mutations are previously unpublished. There are the A5874G mutation in tRNATyr gene, the G12294A mutation in gene, the G4810A mutation in ND2 gene, the T11232C mutation in ND4 gene and the G15723A in cytochrome b gene. The A5874G and G4810A mutations are the first mutations in the tRNATyr and ND2 gene, respectively, to be described in association with human diseases. All but the G4810A mutation were associated with sporadic myopathy without involvement of other tissues. For these sporadic cases, mutant mtDNA was present in only skeletal muscle was not detected in other tissues or in the blood mtDNA of the maternal relatives examined. These data suggested that these mutations might be somatic. Four MELAS cases were found to harbour the G13513A mutation in the ND5 gene previously reported in a single case. Our data indicate that the G13513A is probably the second commonest cause of MELAS in the United Kingdom. Furthermore the phenotypic diversity associated with the G13513A mutation has been extended to include MELAS overlap with Leber's hereditary optic neuropathy. Despite complete sequencing of the entire muscle mtDNA in the nineteen remaining patients, pathogenic mtDNA mutations were not identified in these patients. These data suggest that these patients might harbour genetic defects in nuclear genes. The role of mtDNA background in influencing the phenotypic expression of the common A3243G MELAS mutation in forty-eight unrelated cases was studied. The results indicated that a polymorphism, A12308G, in the tRNALeu(CUN) gene may increase risk of developing stroke in the A3243G patients (relative risk = 2.17). The role of mtDNA background on clinical expression of mtDNA mutations was further observed in association with a 7472C-insertion mutation in tRNASer(UCN) gene. The author described two new unrelated families harbouring the 7472C-insertion which the first family had myoclonic epilepsy and sensorineural hearing loss similar to previous reports. However, the second case had isolated myopathy, in which she had markedly different muscle histochemical findings from other reported cases The intragenic polymorphism, T7472G, in the gene was observed in the second case and might be influence in the phenotypic expression of the 7472C-insertion. In conclusion, this study has defined the molecular genetic basis of mitochondrial diseases in eleven patients. Five novel mtDNA mutations are described and characterised. Evidence is presented that the G13513A mutation is an important cause of the MELAS phenotype. It is suggested that nuclear gene defects probably account for the disease in the remaining nineteen patients. Evidence is presented that the mtDNA haplotype may influence the expression of primary mtDNA defects