title: Mitochondria in cultured human muscle cells depleted of mitochondrial DNA.
abstract: Cultured human muscle cells were depleted of mitochondrial DNA (mtDNA) by prolonged treatment with ethidium bromide (EB). In these respiration-deficient muscle cells neither cytochrome c oxidase activity nor mtDNA were detectable. However, mitochondrial matrix enzymes remained present and were localized in mitochondria-like organelles, as shown by subcellular fractionation. Metabolic labeling showed synthesis of cytochrome c oxidase subunits coded by nuclear DNA (nDNA). These results indicate that depletion of mtDNA in cultured human myoblasts does not inhibit expression of nDNA-coded mitochondrial proteins. The characteristic thread-like pattern of mitochondria was lost in mtDNA-depleted myoblasts, as shown by immunofluorescence with antibodies against cytochrome c oxidase and the F1 part of the mitochondrial ATP synthase (F1-ATPase) and by fluorescence of the carbocyanine dye, 3,3'-dipentyloxacarbocyanine iodide (DiOC5(3)). The organelles visualized by these methods were round and swollen and had a localization different from lysosomes as shown by double-labeling with mitochondrial and lysosomal antibodies. These results indicate that not only synthesis, but also import of mitochondrial proteins into mitochondria-like organelles remains possible in respiration-deficient cells.
authors.affiliation: Department of Neurology, University of Amsterdam, The Netherlands.
authors.forename: N H
authors.initials: NH
authors.lastname: Herzberg
authors.forename: E
authors.initials: E
authors.lastname: Middelkoop
authors.forename: M
authors.initials: M
authors.lastname: Adorf
authors.forename: H L
authors.initials: HL
authors.lastname: Dekker
authors.forename: M J
authors.initials: MJ
authors.lastname: Van Galen
authors.forename: M
authors.initials: M
authors.lastname: Van den Berg
authors.forename: P A
authors.initials: PA
authors.lastname: Bolhuis
authors.forename: C
authors.initials: C
authors.lastname: Van den Bogert
mesh_terms: D002467:Cell Nucleus; D002950:Citrate (si)-Synthase / Q000096:biosynthesis / Q000235:genetics; D004249:DNA Damage; D004272:DNA, Mitochondrial / Q000187:drug effects / Q000235:genetics*; D004579:Electron Transport; D003576:Electron Transport Complex IV / Q000096:biosynthesis / Q000235:genetics; D004996:Ethidium / Q000494:pharmacology; D005786:Gene Expression Regulation; D005969:Glutamate Dehydrogenase / Q000096:biosynthesis / Q000235:genetics; D006801:Humans; D007770:L-Lactate Dehydrogenase / Q000096:biosynthesis / Q000235:genetics; D008931:Mitochondria, Muscle / Q000737:chemistry / Q000648:ultrastructure*; D009124:Muscle Proteins / Q000096:biosynthesis / Q000235:genetics; D010101:Oxygen Consumption; D006180:Proton-Translocating ATPases / Q000096:biosynthesis / Q000235:genetics; D001619:beta-N-Acetylhexosaminidases / Q000096:biosynthesis / Q000235:genetics
chemical_list: D004272:DNA, Mitochondrial; D009124:Muscle Proteins; D007770:L-Lactate Dehydrogenase; D005969:Glutamate Dehydrogenase; D003576:Electron Transport Complex IV; D002950:Citrate (si)-Synthase; D001619:beta-N-Acetylhexosaminidases; D006180:Proton-Translocating ATPases; D004996:Ethidium