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The Nucleotide Sequence, DNA Damage Location, and Protein Stoichiometry Influence the Base Excision Repair Outcome at CAG/CTG Repeats
journal contribution
posted on 2015-12-16, 21:21 authored by Agathi-Vasiliki Goula, Christopher
E. Pearson, Julie Della Maria, Yvon Trottier, Alan E. Tomkinson, David M. Wilson, Karine MerienneExpansion 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.
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Keywords
results showcerebellar stoichiometryBER stoichiometrycerebellummultinucleotide synthesisrepair outcomeinstabilityAPEnucleotide sequenceDNA damage positionProtein Stoichiometry InfluenceLIG 1. DamageDNA Damage Locationhairpin formationmyotonic dystrophyabasic siteCTGcontrol DNA sequencesHD mouse striatumprotein stoichiometryOligonucleotide substratesFENAP lesionrepair efficiencybase excision repairCAGrepair assaysprotein extractsNucleotide Sequencestriatal conditionsBase Excision Repair Outcomecontrol sequencesreconstituted BER protein stoichiometries
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