Heteroplasmy is ubiquitous and stable at the single cell level

Correspondence: [email protected] Department of Oncological Sciences, Icahn School of Medicine at Mount Sinai , One Gustave L. Levy place, New York, 10029, USA Full list of author information is available at the end of the article Abstract Eukaryotic cells carry two genomes, nuclear (nDNA) and mitochondrial (mtDNA), which are ostensibly decoupled in their replication, segregation and inheritance. It is increasingly appreciated that heteroplasmy, the occurrence of multiple mtDNA haplotypes in a cell, plays an important biological role, but its features are not well understood. Until now, accurately determining the diversity of mtDNA has been difficult due to the relatively small amount of mtDNA in each cell (< 1% of the total DNA), the intercellular variability of mtDNA content and copies of mtDNA pseudogenes in nDNA. To understand the nature of heteroplasmy, we developed Mseek, a novel technique that purifies and sequences mtDNA. Mseek yields high purity (> 98%) mtDNA and its ability to detect rare variants is limited only by sequencing depth, providing unprecedented sensitivity and specificity. Using Mseek, we confirmed the ubiquity of heteroplasmy by analyzing mtDNA from a diverse set of cell lines and human samples. Applying Mseek to colonies derived from single cells, we find heteroplasmy is stably maintained in individual daughter cells over multiple cell divisions. Our simulations suggest the stability of heteroplasmy is facilitated by the exchange of mtDNA between cells. We also explicitly demonstrate this exchange by co-culturing cell lines with distinct mtDNA haplotypes. Our results shed new light on the maintenance of heteroplasmy and provide a novel platform to investigate various features of heteroplasmy in normal and diseased tissues.