Bioenergetic consequences of accumulating the common 4977‐bp mitochondrial DNA deletion

Mutations and deletions in mitochondrial DNA (mtDNA) lead to a number of human diseases characterized by neuromuscular degeneration. Accumulation of truncated mtDNA molecules (Δ‐mtDNA) lacking a specific 4977‐bp fragment, the common deletion, leads to three related mtDNA diseases : Pearson's syndrome; Kearns‐Sayre syndrome; and chronic progressive external ophthalmoplegia (CPEO). In addition, the proportion of Δ‐mtDNA present increases with age in a range of tissues. Consequently, there is considerable interest in the effects of the accumulation of Δ‐mtDNA on cell function. The 4977‐bp deletion affects genes encoding 7 polypeptide components of the mitochondrial respiratory chain, and 5 of the 22 tRNAs necessary for mitochondrial protein synthesis. To determine how the accumulation of Δ‐mtDNA affects oxidative phosphorylation we constructed a series of cybrids by fusing a human osteosarcoma cell line depleted of mtDNA (@gR0) with enucleated skin fibroblasts from a CPEO patient. The ensuing cybrids contained 0−86 %Δ‐mtDNA and all had volumes, protein contents, plasma‐membrane potentials and mitochondrial contents similar to those of the parental cell line. The bioenergetic consequences of accumulating Δ‐mtDNA were assessed by measuring the mitochondrial membrane potential, rate of ATP synthesis and ATP/ADP ratio. In cybrids containing less than 50−55 %Δ‐mtDNA, these bioenergetic functions were equivalent to those of cybrids with intact mtDNA. However, once the proportion of Δ‐mtDNA exceeded this threshold, the mitochondrial membrane potential, rate of ATP synthesis, and cellular ATP/ADP ratio decreased. These bioenergetic deficits will contribute to the cellular pathology associated with the accumulation of Δ‐mtDNA in the target tissues of patients with mtDNA diseases.