Dataset Information

The nucleotide sequence, DNA damage location, and protein stoichiometry influence the base excision repair outcome at CAG/CTG repeats.

ABSTRACT: Expansion of CAG/CTG repeats is the underlying cause of >14 genetic disorders, including Huntington's disease (HD) and myotonic dystrophy. The mutational process is ongoing, with increases in repeat size enhancing the toxicity of the expansion in specific tissues. In many repeat diseases, the repeats exhibit high instability in the striatum, whereas instability is minimal in the cerebellum. We provide molecular insights into how base excision repair (BER) protein stoichiometry may contribute to the tissue-selective instability of CAG/CTG repeats by using specific repair assays. Oligonucleotide substrates with an abasic site were mixed with either reconstituted BER protein stoichiometries mimicking the levels present in HD mouse striatum or cerebellum, or with protein extracts prepared from HD mouse striatum or cerebellum. In both cases, the repair efficiency at CAG/CTG repeats and at control DNA sequences was markedly reduced under the striatal conditions, likely because of the lower level of APE1, FEN1, and LIG1. Damage located toward the 5' end of the repeat tract was poorly repaired, with the accumulation of incompletely processed intermediates as compared to an AP lesion in the center or at the 3' end of the repeats or within control sequences. Moreover, repair of lesions at the 5' end of CAG or CTG repeats involved multinucleotide synthesis, particularly at the cerebellar stoichiometry, suggesting that long-patch BER processes lesions at sequences susceptible to hairpin formation. Our results show that the BER stoichiometry, nucleotide sequence, and DNA damage position modulate repair outcome and suggest that a suboptimal long-patch BER activity promotes CAG/CTG repeat instability.

SUBMITTER: Goula AV

PROVIDER: S-EPMC3357312 | biostudies-literature | 2012 May

REPOSITORIES: biostudies-literature

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The nucleotide sequence, DNA damage location, and protein stoichiometry influence the base excision repair outcome at CAG/CTG repeats.

Goula Agathi-Vasiliki AV Pearson Christopher E CE Della Maria Julie J Trottier Yvon Y Tomkinson Alan E AE Wilson David M DM Merienne Karine K

Biochemistry 20120423 18

Expansion of CAG/CTG repeats is the underlying cause of >14 genetic disorders, including Huntington's disease (HD) and myotonic dystrophy. The mutational process is ongoing, with increases in repeat size enhancing the toxicity of the expansion in specific tissues. In many repeat diseases, the repeats exhibit high instability in the striatum, whereas instability is minimal in the cerebellum. We provide molecular insights into how base excision repair (BER) protein stoichiometry may contribute to ...[more]

PMID: 22497302

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Project description:Huntington's disease (HD) is a progressive neurodegenerative disorder caused by expansion of an unstable CAG repeat in the coding sequence of the Huntingtin (HTT) gene. Instability affects both germline and somatic cells. Somatic instability increases with age and is tissue-specific. In particular, the CAG repeat sequence in the striatum, the brain region that preferentially degenerates in HD, is highly unstable, whereas it is rather stable in the disease-spared cerebellum. The mechanisms underlying the age-dependence and tissue-specificity of somatic CAG instability remain obscure. Recent studies have suggested that DNA oxidation and OGG1, a glycosylase involved in the repair of 8-oxoguanine lesions, contribute to this process. We show that in HD mice oxidative DNA damage abnormally accumulates at CAG repeats in a length-dependent, but age- and tissue-independent manner, indicating that oxidative DNA damage alone is not sufficient to trigger somatic instability. Protein levels and activities of major base excision repair (BER) enzymes were compared between striatum and cerebellum of HD mice. Strikingly, 5'-flap endonuclease activity was much lower in the striatum than in the cerebellum of HD mice. Accordingly, Flap Endonuclease-1 (FEN1), the main enzyme responsible for 5'-flap endonuclease activity, and the BER cofactor HMGB1, both of which participate in long-patch BER (LP-BER), were also significantly lower in the striatum compared to the cerebellum. Finally, chromatin immunoprecipitation experiments revealed that POLbeta was specifically enriched at CAG expansions in the striatum, but not in the cerebellum of HD mice. These in vivo data fit a model in which POLbeta strand displacement activity during LP-BER promotes the formation of stable 5'-flap structures at CAG repeats representing pre-expanded intermediate structures, which are not efficiently removed when FEN1 activity is constitutively low. We propose that the stoichiometry of BER enzymes is one critical factor underlying the tissue selectivity of somatic CAG expansion.

Dataset Information

The nucleotide sequence, DNA damage location, and protein stoichiometry influence the base excision repair outcome at CAG/CTG repeats.

Publications

The nucleotide sequence, DNA damage location, and protein stoichiometry influence the base excision repair outcome at CAG/CTG repeats.

Similar Datasets

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