Ageing muscle: clonal expansions of mitochondrial DNA point mutations and deletions cause focal impairment of mitochondrial function.
Although mitochondrial DNA deletions have been shown to accumulate in cytochrome c oxidase deficient muscle fibres of ageing muscle, this has not been demonstrated for point mutations. In this study, we investigated the occurrence of mitochondrial DNA alterations (point mutations and deletions) in cytochrome c oxidase deficient muscle fibres from 14 individuals, without muscle disease, aged 69-82 years. Immunohistochemical investigation showed that the majority of the cytochrome c oxidase deficient muscle fibres expressed reduced levels of subunit II of cytochrome c oxidase, which is encoded by mitochondrial DNA, whereas there was normal or increased expression of subunit IV of cytochrome c oxidase, which is encoded by nuclear DNA. This pattern is typical for mitochondrial DNA mutations causing impaired mitochondrial translation. Single muscle fibres (109 cytochrome c oxidase deficient and 109 normal fibres) were dissected and their DNA extracted. Mitochondrial DNA point mutations were searched for in five tRNA genes by denaturing gradient gel electrophoresis while deletions were looked for by polymerase chain reaction amplification. High levels of clonally expanded point mutations were identified in eight cytochrome c oxidase deficient fibres but in none of the normal ones. They included the previously described pathogenic tRNALeu(UUR)A3243G and tRNALysA8344G mutations and three original mutations: tRNAMetT4460C, tRNAMetG4421A, and a 3-bp deletion in the tRNALeu(UUR) gene. Four different large-scale mitochondrial DNA deletions were identified in seven cytochrome c oxidase deficient fibres and in one of the normal ones. There was no evidence of depletion of mitochondrial DNA by in situ hybridisation experiments. Our data show that mitochondrial DNA point mutations, as well as large-scale deletions, are associated with cytochrome c oxidase deficient muscle fibre segments in ageing. Their focal accumulation causes significant impairment of mitochondrial function in individual cells in spite of low overall levels of mitochondrial DNA mutations in muscle.
Fayet
Guillemette
G
Inserm U523, Institut de Myologie, Hôpital Pitié-Salpêtrière, Paris, France.
Jansson
Monica
M
Sternberg
Damien
D
Moslemi
Ali Reza
AR
Blondy
Patricia
P
Lombès
Anne
A
Fardeau
Michel
M
Oldfors
Anders
A
D000368:Aged; D000369:Aged, 80 and over; D000375:Aging / Q000502:physiology*; D001483:Base Sequence / Q000235:genetics; D030401:Cytochrome-c Oxidase Deficiency / Q000235:genetics*; D004272:DNA, Mitochondrial / Q000235:genetics*; D003576:Electron Transport Complex IV / Q000378:metabolism; D005260:Female; D017353:Gene Deletion; D006801:Humans; D007527:Isoenzymes / Q000378:metabolism; D008297:Male; D008931:Mitochondria, Muscle / Q000502:physiology; D028361:Mitochondrial Diseases / Q000235:genetics*; D018485:Muscle Fibers, Skeletal / Q000502:physiology; D018482:Muscle, Skeletal / Q000201:enzymology*; D017354:Point Mutation / Q000502:physiology*
D004272:DNA, Mitochondrial; D007527:Isoenzymes; D003576:Electron Transport Complex IV