Mechanisms of Human Mitochondrial DNA Maintenance: The Determining Role of Primary Sequence and Length over Function

Abstract

Although the regulation of mitochondrial DNA (mtDNA) copy number is performed by nuclear-coded factors, very little is known about the mechanisms controlling this process. We attempted to introduce nonhuman ape mtDNA into human cells harboring either no mtDNA or mutated mtDNAs (partial deletion and tRNA gene point mutation). Unexpectedly, only cells containing no mtDNA could be repopulated with nonhuman ape mtDNA. Cells containing a defective human mtDNA did not incorporate or maintain ape mtDNA and therefore died under selection for oxidative phosphorylation function. On the other hand, foreign human mtDNA was readily incorporated and maintained in these cells. The suicidal preference for self-mtDNA showed that functional parameters associated with oxidative phosphorylation are less relevant to mtDNA maintenance and copy number control than recognition of mtDNA self-determinants. Non-self-mtDNA could not be maintained into cells with mtDNA even if no selection for oxidative phosphorylation was applied. The repopulation kinetics of several mtDNA forms after severe depletion by ethidium bromide treatment showed that replication and maintenance of mtDNA in human cells are highly dependent on molecular features, because partially deleted mtDNA molecules repopulated cells significantly faster than full-length mtDNA. Taken together, our results suggest that mtDNA copy number may be controlled by competition for limiting levels of trans-acting factors that recognize primarily mtDNA molecular features. In agreement with this hypothesis, marked variations in mtDNA levels did not affect the transcription of nuclear-coded factors involved in mtDNA replication.

Conflicting levels of selection in the accumulation of mitochondrial defects in Saccharomyces cerevisiae

“…A within-cell advantage for mutant mitochondrial genomes has been suggested by the results of experiments with mice (11) and with cultures of human cells (7,9,10,13). In some experiments, this within-cell advantage has been tissue-specific, appearing in one tissue or cell type but being absent or reversed in another (11).…”

“…In some experiments, this within-cell advantage has been tissue-specific, appearing in one tissue or cell type but being absent or reversed in another (11). This pattern has been interpreted as an indication that the replication of mtDNA is regulated by one or more factors encoded by the nucleus that are expressed in a tissue-specific way and that discriminate among genetically different mitochondrial chromosomes (13). Alternatively, our model predicts that such tissue-specific differences may result from differences in the general properties of cells (14), or even the culture conditions, that would alter the relative intensities of within-and among-cell selection.…”

“…Mitochondria also lack some of the DNA repair pathways that operate in the nucleus to reduce mutation rates (6). However, some defects may accumulate because mutant mitochondrial chromosomes have a fitness advantage over functional mitochondrial genomes within cells (7)(8)(9)(10)(11)(12)(13)(14). The opportunity for natural selection to occur within cells stems from the fact that each cell contains multiple mitochondrial genomes that replicate independently of the cell.…”

Loss of mutant mitochondrial DNA harboring the MELAS A3243G mutation in human cybrid cells after cell-cell fusion with normal tissue-derived fibroblast cells

“…It was reported that human mtDNA is preferentially maintained over ape mtDNA in human cybrid and hybrid cells (17). The report indicated that the human nucleus had a strong preference for human mtDNA rather than ape normal mtDNA, even if pathogenic mutations exist in the human mtDNA.…”

Inter-Species Therapeutic Cloning: The Looming Problem of Mitochondrial DNA and Two Possible Solutions

“…Such inefficiency has been shown in cytoplasmic hybrid cells using even very closely-related species. 6 Unfortunately, two recent papers 2,3 appear to dash these hopes. Careful monitoring of the relative and absolute abundance of donor-and oocyte-derived mtDNA in the early SCNT-derived embryo as it develops to the blastocyst stage shows that, when the donor is a primate and the oocyte is from either cow or rabbit, the donor mtDNA is not merely out-replicated but is actually lost in the few cell divisions following the initiation of mitochondrial biogenesis.…”