On the timing and the extent of clonal expansion of mtDNA deletions: Evidence from single-molecule PCR

mtDNA deletions are pathogenic mutations that remove substantial portions of the mitochondrial genome. mtDNA deletions accumulate with age and have been implicated in various degenerative diseases. There are multiple mtDNA per cell and mtDNA mutations become toxic only if they accumulate to substantial intracellular levels, i.e. exceed so-called “phenotypic threshold”. This is usually achieved via clonal expansion of a single initial mutated molecule. Intracellular mitochondrial genomes are analogous to a population of individuals in that mitochondria are born by division and die by degradation. Clonal expansion within cells is thus analogous to genetic drift within populations and is driven by a combination of random processes and selection. mtDNA mutations occurring early in development are expected to end up spread across tissues, while mutations of late origin are expected to be localized, i.e. limited a single post-mitotic cell or progeny of a single stem cell. We have explored the extent and timing of clonality of mtDNA deletion in human muscle using single-molecule PCR. We analyzed deletions from two nearby locations within the same tissue sample. Altogether we analyzed over 130 mutant molecules, but almost every deletion type detected was represented by several identical mutant molecules, so that altogether there were only 21 different kinds of deletions, implying that essentially all deletions were clonal. At the same time the sets of deletions in the two locations were completely different. This observation implies that all of the clonal expansions spanned very small areas and therefore that the corresponding mutations were likely events of older age. More studies are necessary to further validate these findings in muscle and to explore the other tissues.