Differentiation of neural stem cells (NSCs) involves the activation of aerobic metabolism, which is dependent on mitochondrial function. Here, we show that the differentiation of NSCs involves robust increases in mitochondrial mass, mitochondrial DNA (mtDNA) copy number, and respiration capacity. The increased respiration activity renders mtDNA vulnerable to oxidative damage, and NSCs defective for the mitochondrial 8-oxoguanine DNA glycosylase (OGG1) function accumulate mtDNA damage during the differentiation. The accumulated mtDNA damages in ogg1 2/2 cells inhibit the normal maturation of mitochondria that is manifested by reduced cellular levels of mitochondrial encoded complex proteins (complex I[cI], cIII, and cIV) with normal levels of the nuclear encoded cII present. The specific cI activity and inner membrane organization of respiratory complexes are similar in wt and ogg1 2/2 cells, inferring that mtDNA damage manifests itself as diminished mitochondrial biogenesis rather than the generation of dysfunctional mitochondria. Aerobic metabolism increases during differentiation in wild-type cells and to a lesser extent in ogg1 2/2 cells, whereas anaerobic rates of metabolism are constant and similar in both cell types. Our results demonstrate that mtDNA integrity is essential for effective mitochondrial maturation during NSC differentiation. STEM CELLS

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Cockayne syndrome (CS) is a severe, complex, progressive and autosomal recessive disease characterized by developmental, skeletal, neurological and muscular failure, as well as a premature aging phenotype [1]. Patients with CS can be assigned to two different complementation groups, with the majority carrying mutations in the CSB gene [2]. A clinically identified CSB truncation (CS1AN) has been genetically introduced into a mouse, which displays similar but milder CS phenotypes [3]. The CSB protein plays distinct roles in different DNA repair pathways. It is involved in the transcriptioncoupled repair of UV-induced DNA damage, which is reflected by the sensitivity of human and murine csb m ⁄ m fibroblasts to UV radiation. Furthermore, CSB is involved in the global repair of different types of endogenously formed DNA damage and also participates in regulation of the major nuclear and mitochondrial 8-oxoguanine (8oxoG) Cockayne syndrome (CS) is a complex, progressive disease that involves neurological and developmental impairment and premature aging. The majority of CS patients have mutations in the CSB gene. The CSB protein is involved in multiple DNA repair pathways and CSB mutated cells are sensitive to a broad spectrum of genotoxic agents. We tested the hypothesis that sensitivity to such genotoxins could be mediated by mitochondrial dysfunction as a consequence of the CSB mutation. mtDNA from csb m ⁄ m mice accumulates oxidative damage including 8-oxoguanine, and cells from this mouse are hypersensitive to the mitochondrial oxidant menadione. Inhibitors of mitochondrial complexes and the glycolysis inhibitor 2-deoxyglucose kill csb m ⁄ m cells more efficiently than wild-type cells, via a mechanism that does not correlate with mtDNA damage formation. Menadione depletes cellular ATP, and recovery after depletion is slower in csb m ⁄ m cells. The bioenergetic alteration in csb m ⁄ m cells parallels the simpler organization of supercomplexes consisting of complexes I, III and IV in addition to partially disassembled complex V in the inner mitochondrial membrane. Exposing wild-type cells to DNA intercalating agents induces complex alterations, suggesting a link between mtDNA integrity, respiratory complexes and mitochondrial function. Thus, mitochondrial dysfunction may play a role in the pathology of CS.Abbreviations