Search

Mutations in genes encoding components of the mitochondrial DNA (mtDNA) replication machinery cause mtDNA depletion syndromes (MDSs), which associate ocular features with severe neurological syndromes. Here, we identified heterozygous missense mutations in single-strand binding protein 1 (SSBP1) in 5 unrelated families, leading to the R38Q and R107Q amino acid changes in the mitochondrial single-stranded DNA-binding protein, a crucial protein involved in mtDNA replication. All affected individuals presented optic atrophy, associated with foveopathy in half of the cases. To uncover the structural features underlying SSBP1 mutations, we determined a revised SSBP1 crystal structure. Structural analysis suggested that both mutations affect dimer interactions and presumably distort the DNA-binding region. Using patient fibroblasts, we validated that the R38Q variant destabilizes SSBP1 dimer/tetramer formation, affects mtDNA replication, and induces mtDNA depletion. Our study showing that mutations in SSBP1 cause a form of dominant optic atrophy frequently accompanied with foveopathy brings insights into mtDNA maintenance disorders., Introduction Mitochondrial diseases are clinically heterogeneous and can be caused by mutations in either nuclear or mitochondrial DNA (mtDNA) (1). Mutations in genes known to be essential for mtDNA maintenance [...]

BackgroundMammals have seven NAD-dependent protein deacetylases. These proteins, called sirtuins, are homologous to yeast Sir2, and are emerging as important regulators of lifespan and intermediary metabolism. Three mammalian sirtuins, SIRT3-5 are mitochondrial. Sirtuins are highly conserved between species, yet mouse SIRT3 was reported to be markedly shorter than its human counterpart and to lack the N-terminal mitochondrial targeting signal present in the human protein.ResultsWe have isolated a novel mouse SIRT3 splice variant. This cDNA contains two translation initiation codons upstream of the originally reported start site. We show, using immunofluorescence and protein expression analysis that these longer variants are expressed and efficiently targeted to mitochondria, and that the processed forms of these longer variants are identical in size to the endogenous mouse SIRT3. We also show that the previously described form of SIRT3 is not mitochondrial.ConclusionsOur observations point to a high level of conservation of SIRT3 as a mitochondrial protein in mice and human and indicate that several previous studies, which addressed mouse Sirt3 function, need to be re-evaluated.

The mitochondrial theory of aging proposes that reactive oxygen species (ROS) generated inside the cell will lead, with time, to increasing amounts of oxidative damage to various cell components. The main site for ROS production is the respiratory chain inside the mitochondria and accumulation of mtDNA mutations, and impaired respiratory chain function have been associated with degenerative diseases and aging. The theory predicts that impaired respiratory chain function will augment ROS production and thereby increase the rate of mtDNA mutation accumulation, which, in turn, will further compromise respiratory chain function. Previously, we reported that mice expressing an error-prone version of the catalytic subunit of mtDNA polymerase accumulate a substantial burden of somatic mtDNA mutations, associated with premature aging phenotypes and reduced lifespan. Here we show that these mtDNA mutator mice accumulate mtDNA mutations in an approximately linear manner. The amount of ROS produced was normal, and no increased sensitivity to oxidative stress-induced cell death was observed in mouse embryonic fibroblasts from mtDNA mutator mice, despite the presence of a severe respiratory chain dysfunction. Expression levels of antioxidant defense enzymes, protein carbonylation levels, and aconitase enzyme activity measurements indicated no or only minor oxidative stress in tissues from mtDNA mutator mice. The premature aging phenotypes in mtDNA mutator mice are thus not generated by a vicious cycle of massively increased oxidative stress accompanied by exponential accumulation of mtDNA mutations. We propose instead that respiratory chain dysfunction per se is the primary inducer of premature aging in mtDNA mutator mice. mitochondria | mtDNA mutator mice

Defects of mitochondrial DNA (mtDNA) maintenance have recently been associated with inherited neurodegenerative and muscle diseases and the aging process. Twinkle is a nuclear-encoded mtDNA helicase, dominant mutations of which cause adult-onset progressive external ophthalmoplegia (PEO) with multiple mtDNA deletions. We have generated transgenic mice expressing mouse Twinkle with PEO patient mutations. Multiple mtDNA deletions accumulate in the tissues of these mice, resulting in progressive respiratory dysfunction and chronic late-onset mitochondrial disease starting at 1 year of age. The muscles of the mice faithfully replicate all of the key histological, genetic, and biochemical features of PEO patients. Furthermore, the mice have progressive deficiency of cytochrome c oxidase in distinct neuronal populations. These 'deletor' mice do not, however, show premature aging, indicating that subtle accumulation of mtDNA deletions and progressive respiratory chain dysfunction are not sufficient to accelerate aging. This model is a valuable tool for therapy development and testing for adult-onset mitochondrial disorders. mouse model | progressive external ophthalmoplegia | mitochondrial DNA replication

Author(s): Aleksandra Trifunovic [1, 2]; Anna Wredenberg [1, 2]; Maria Falkenberg [1]; Johannes N. Spelbrink [3]; Anja T. Rovio [3]; Carl E. Bruder [4]; Mohammad Bohlooly-Y [4]; Sebastian Gidlöf [1, [...]

SUPV3L1 encodes a helicase that is mainly localized in the mitochondria. It has been shown in vitro to possess both double‐stranded RNA and DNA unwinding activity that is ATP‐dependent. Here we report the first two patients for this gene who presented with a homozygous preliminary stop codon resulting in a C‐terminal truncation of the SUPV3L1 protein. They presented with a characteristic phenotype of neurodegenerative nature with progressive spastic paraparesis, growth restriction, hypopigmentation, and predisposition to autoimmune disease. Ophthalmological examination showed severe photophobia with corneal erosions, optic atrophy, and pigmentary retinopathy, while neuroimaging showed atrophy of the optic chiasm and the pons with calcification of putamina, with intermittent and mild elevation of lactate. We show that the amino acids that are eliminated by the preliminary stop codon are highly conserved and are predicted to form an amphipathic helix. To investigate if the mutation causes mitochondrial dysfunction, we examined fibroblasts of the proband. We observed very low expression of the truncated protein, a reduction in the mature ND6 mRNA species as well as the accumulation of double‐stranded RNA. Lentiviral complementation with the full‐length SUPV3L1 cDNA partly restored the observed RNA phenotypes, supporting that the SUPV3L1 mutation in these patients is pathogenic and the cause of the disease. [ABSTRACT FROM AUTHOR]