Mitochondrial DNA spectra of single human CD34+ cells, T cells, B cells, and granulocytes

Abstract

AbstractPreviously, we described the age-dependent accumulation of mitochondrial DNA (mtDNA) mutations, leading to a high degree of mtDNA heterogeneity among normal marrow and blood CD34+ clones and in granulocytes. We established a method for sequence analysis of single cells. We show marked, distinct mtDNA heterogeneity from corresponding aggregate sequences in isolated cells of 5 healthy adult donors—37.9% ± 3.6% heterogeneity in circulating CD34+ cells, 36.4% ± 14.1% in T cells, 36.0% ± 10.7% in B cells, and 47.7% ± 7.4% in granulocytes. Most heterogeneity was caused by poly-C tract variability; however, base substitutions were also prevalent, as follows: 14.7% ± 5.7% in CD34+ cells, 15.2% ± 9.0% in T cells, 15.4% ± 6.7% in B cells, and 32.3% ± 2.4% in granulocytes. Many poly-C tract length differences and specific point mutations seen in these same donors but assayed 2 years earlier were still present in the new CD34+ samples. Additionally, specific poly-C tract differences and point mutations were frequently shared among cells of the lymphoid and myeloid lineages. Secular stability and lineage sharing of mtDNA sequence variability suggest that mutations arise in the lymphohematopoietic stem cell compartment and that these changes may be used as a natural genetic marker to estimate the number of active stem cells.