Prominent mitochondrial DNA recombination intermediates in human heart muscle

Abstract

Recombination intermediates containing four-way (Holliday) junctions are generated during DNA repair and replication in many systems, including yeast mitochondrial DNA (mtDNA). In contrast, convincing evidence for recombination in mammalian mtDNA is lacking. We have used two-dimensional agarose-gel electrophoresis to analyse non-linear forms of mtDNA in human heart muscle. Replication intermediates from both the coupled and strand-asynchronous mtDNA replication pathways were detected. An additional class of non-linear molecules, with the electrophoretic properties of four-way junctions, was also prominent. These molecules were insensitive to topoisomerase I or RNase H, but were diminished by branch migration or RuvC treatment. Junctional molecules were detected in all regions of the mitochondrial genome, were found in myocardial DNA from young and old adults, but were present at lower levels in skeletal muscle and placenta. We suggest that they could represent intermediates of mtDNA repair, given their prevalence in the oxyradical-rich environment of heart muscle mitochondria.

Mitochondrial Genome Maintenance: Roles for Nuclear Nonhomologous End-Joining Proteins inSaccharomyces cerevisiae

“…However, more direct experimental evidence for recombination and end joining in mammalian mitochondria, derived from in vivo and in vitro studies, has accumulated in recent years (Poulton et al 1993;Thyagarajan et al 1996;Coffey et al 1999;Lakshmipathy and Campell 1999;Kajander et al 2001;Kraytsberg et al 2004). The finding that some of the known nuclear DSBR proteins are localized to the mitochondria has suggested that, like BER, some of these pathways may be operational in both nuclear and mitochondrial compartments.…”

“…Interestingly, the majority of mtDNA deletions are flanked by short repetitive sequences, implicating recombination as a possible mechanism (Bianchi et al 2001;Krishnan et al 2008). Recombination of the mitochondrial genome has been well accepted in yeast and plants; and recombinant mtDNA molecules and homologous recombination (HR) activity have been detected in mammalian cells in vitro and in vivo (Poulton et al 1993;Thyagarajan et al 1996;Lakshmipathy and Campell 1999;Kajander et al 2001;Kraytsberg et al 2004). These studies and others provide evidence for mitochondrial double-strand break repair (mtDSBR); however, no proteins have been identified that function in these pathways.…”

Mechanism of Homologous Recombination and Implications for Aging-Related Deletions in Mitochondrial DNA

“…By using two-dimensional agarose gel electrophoresis, an early report showed that molecular species suggestive of Holliday junctions exist in the human heart mtDNA (86). This observation has now been further corroborated by transmission electron microscopy (87).…”

The mitochondrial theory of aging: dead or alive?

“…The replication and turnover of mtDNA continue even in cells that are irreversibly committed to a differentiated fate, and which will never again re-enter the cell cycle. This is abundantly clear from the fact that mtDNA replication intermediates can be easily detected, electrophoretically, in terminally differentiated tissues such as cardiac and skeletal muscle (Kajander et al ., 2001). Therefore, somatic mutation of mtDNA need not be a one-way street.…”