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Terminally differentiated muscle cells are defective in base excision DNA repair and hypersensitive to oxygen injury.

ABSTRACT: The differentiation of skeletal myoblasts is characterized by permanent withdrawal from the cell cycle and fusion into multinucleated myotubes. Muscle cell survival is critically dependent on the ability of cells to respond to oxidative stress. Base excision repair (BER) is the main repair mechanism of oxidative DNA damage. In this study, we compared the levels of endogenous oxidative DNA damage and BER capacity of mouse proliferating myoblasts and their differentiated counterpart, the myotubes. Changes in the expression of oxidative stress marker genes during differentiation, together with an increase in 8-hydroxyguanine DNA levels in terminally differentiated cells, suggested that reactive oxygen species are produced during this process. The repair of 2-deoxyribonolactone, which is exclusively processed by long-patch BER, was impaired in cell extracts from myotubes. The repair of a natural abasic site (a preferred substrate for short-patch BER) also was delayed. The defect in BER of terminally differentiated muscle cells was ascribed to the nearly complete lack of DNA ligase I and to the strong down-regulation of XRCC1 with subsequent destabilization of DNA ligase IIIalpha. The attenuation of BER in myotubes was associated with significant accumulation of DNA damage as detected by increased DNA single-strand breaks and phosphorylated H2AX nuclear foci upon exposure to hydrogen peroxide. We propose that in skeletal muscle exacerbated by free radical injury, the accumulation of DNA repair intermediates, due to attenuated BER, might contribute to myofiber degeneration as seen in sarcopenia and many muscle disorders.

SUBMITTER: Narciso L

PROVIDER: S-EPMC2040456 | biostudies-literature | 2007 Oct

REPOSITORIES: biostudies-literature

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Terminally differentiated muscle cells are defective in base excision DNA repair and hypersensitive to oxygen injury.

Narciso Laura L Fortini Paola P Pajalunga Deborah D Franchitto Annapaola A Liu Pingfang P Degan Paolo P Frechet Mathilde M Demple Bruce B Crescenzi Marco M Dogliotti Eugenia E

Proceedings of the National Academy of Sciences of the United States of America 20071016 43

The differentiation of skeletal myoblasts is characterized by permanent withdrawal from the cell cycle and fusion into multinucleated myotubes. Muscle cell survival is critically dependent on the ability of cells to respond to oxidative stress. Base excision repair (BER) is the main repair mechanism of oxidative DNA damage. In this study, we compared the levels of endogenous oxidative DNA damage and BER capacity of mouse proliferating myoblasts and their differentiated counterpart, the myotubes. ...[more]

PMID: 17940040

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Project description:BACKGROUND:Atherosclerotic plaques demonstrate extensive accumulation of oxidative DNA damage, predominantly as 8-oxoguanine (8oxoG) lesions. 8oxoG is repaired by base excision repair enzymes; however, the mechanisms regulating 8oxoG accumulation in vascular smooth muscle cells (VSMCs) and its effects on their function and in atherosclerosis are unknown. METHODS:We studied levels of 8oxoG and its regulatory enzymes in human atherosclerosis, the mechanisms regulating 8oxoG repair and the base excision repair enzyme 8oxoG DNA glycosylase I (OGG1) in VSMCs in vitro, and the effects of reducing 8oxoG in VSMCs in atherosclerosis in ApoE-/- mice. RESULTS:Human plaque VSMCs showed defective nuclear 8oxoG repair, associated with reduced acetylation of OGG1. OGG1 was a key regulatory enzyme of 8oxoG repair in VSMCs, and its acetylation was crucial to its repair function through regulation of protein stability and expression. p300 and sirtuin 1 were identified as the OGG1 acetyltransferase and deacetylase regulators, respectively, and both proteins interacted with OGG1 and regulated OGG1 acetylation at endogenous levels. However, p300 levels were decreased in human plaque VSMCs and in response to oxidative stress, suggesting that reactive oxygen species-induced regulation of OGG1 acetylation could be caused by reactive oxygen species-induced decrease in p300 expression. We generated mice that express VSMC-restricted OGG1 or an acetylation defective version (SM22?-OGG1 and SM22?-OGG1K-R mice) and crossed them with ApoE-/- mice. We also studied ApoE-/- mice deficient in OGG1 (OGG1-/-). OGG1-/- mice showed increased 8oxoG in vivo and increased atherosclerosis, whereas mice expressing VSMC-specific OGG1 but not the acetylation mutant OGG1K-R showed markedly reduced intracellular 8oxoG and reduced atherosclerosis. VSMC OGG1 reduced telomere 8oxoG accumulation, DNA strand breaks, cell death and senescence after oxidant stress, and activation of proinflammatory pathways. CONCLUSIONS:We identify defective 8oxoG base excision repair in human atherosclerotic plaque VSMCs, OGG1 as a major 8oxoG repair enzyme in VSMCs, and p300/sirtuin 1 as major regulators of OGG1 through acetylation/deacetylation. Reducing oxidative damage by rescuing OGG1 activity reduces plaque development, indicating the detrimental effects of 8oxoG on VSMC function.

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Terminally differentiated muscle cells are defective in base excision DNA repair and hypersensitive to oxygen injury.

Publications

Terminally differentiated muscle cells are defective in base excision DNA repair and hypersensitive to oxygen injury.

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