Lipid and DNA peroxidation give rise to DNA adducts, the most abundant of which is 3-(2-deoxy-β-D-erythro-pentofuranosyl)pyrimido[1,2-α]purin-10(3H)-one (M1dG). While previous studies have reported higher levels of M1dG in mitochondrial versus genomic DNA, perhaps not surprisingly given the relative dearth of repair pathways in the mitochondrion compared to the nucleus, little research effort has been dedicated to understanding mitochondrial M1dG formation or its potential role in disease pathogenesis. Herein we present data confirming that levels of M1dG are indeed higher basally in the mitochondrion than in the nucleus, and that these levels are elevated upon exogenous electrophile stimulation in multiple human cell lines. Notably, mitochondrial levels were particularly high in RAW264.7 macrophages, leading us to hypothesize a correlation between oxidative stress and mitochondrial M1dG levels. To test this theory, cells were treated with rotenone, which inhibits Complex I of the electron transport chain. Rotenone treatment led to increased levels of M1dG, an increase that could be reversed by the concomitant addition of a mitochondria-targeted free radical scavenger mitoTEMPO. Furthermore, mitoTEMPO treatment reduced basal mitochondrial M1dG levels by about two-fold, while the ubiquitously distributed free radical scavenger TEMPOL had no effect on either untreated or rotenone-challenged cells. To assess whether this observed correlation between oxidative stress and mitochondrial M1dG levels applied in an animal model of disease, we isolated pulmonary microvascular endothelial cells from transgenic mice bearing a mutant BMPR2 gene, which predisposes them to develop pulmonary arterial hypertension, a reported mitochondria-linked disease state. Indeed, levels of mitochondrial M1dG were elevated in the BMPR2 mutant mice compared to their wild-type counterparts. Collectively, our data provide evidence of a strong correlation between oxidative stress and mitochondrial M1dG levels and suggest that this adduct may be an ideal biomarker for monitoring disease progression in conditions characterized by mitochondrial dysfunction.

Mitochondrial M1dG levels linked to oxidative stress and mitochondrial dysfunction in disease

254th National Meeting (2017)

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