Abstract
Oxidative DNA damage and base excision repair (BER) play important roles in modulating trinucleotide repeat (TNR) instability that is associated with human neurodegenerative diseases and cancer. We have reported that BER of base lesions can lead to TNR instability. However, it is unknown if modifications of the sugar in an abasic lesion modulate TNR instability. In this study, we characterized the effects of the oxidized sugar, 5’-(2-phosphoryl-1,4-dioxobutane)(DOB) in CAG repeat tracts on the activities of key BER enzymes, as well as on repeat instability. We found that DOB crosslinked with DNA polymerase β and inhibited its synthesis activity in CAG repeat tracts. Surprisingly, we found that DOB also formed crosslinks with DNA ligase I and inhibited its ligation activity, thereby reducing the efficiency of BER. This subsequently resulted in the accumulation of DNA strand breaks in a CAG repeat tract. Our study provides important new insights into the adverse effects of an oxidized abasic lesion on BER and suggests a potential alternate repair pathway through which an oxidized abasic lesion may modulate TNR instability.
Highlights
Trinucleotide repeat (TNR) expansions are associated with over 40 human neurodegenerative diseases, including Huntington’s disease (CAG/CTG), Friedreich’s ataxia (GAA/TTC), fragile X syndrome (CGG/CCG), and myotonic dystrophy (CTG/containing a nick with a downstream (CAG)) [1, 2]
We asked whether this crosslink affects Base excision repair (BER) efficiency and TNR instability when the DOB lesion is present in a TNR tract
Since the DOB lesion is an oxidized sugar, which is refractory to pol β dRP lyase activity due to the fact that only a native sugar, dRP [36] can be directly removed by the dRP lyase activity via β-elimination [37] during short-patch BER, the lesion can only be removed through the long-patch BER subpathway, during which pol β performs multi-nucleotide gap-filling synthesis and/or strand displacement synthesis and coordinates with flap endonuclease 1 (FEN1) removal of a 5’-flap that contains 5’-DOB
Summary
Trinucleotide repeat (TNR) expansions are associated with over 40 human neurodegenerative diseases, including Huntington’s disease (CAG/CTG), Friedreich’s ataxia (GAA/TTC), fragile X syndrome (CGG/CCG), and myotonic dystrophy (CTG/CAG) [1, 2]. TNR instability results from the formation of secondary structures such as hairpins, triplexes, quadruplexes, and sticky DNA [3,4,5] during DNA replication, repair, recombination, and gene transcription [3, 6,7,8]. Base excision repair (BER) within TNR tracts plays a critical role in modulating repeat instability [9,10,11], which results from a loss of coordination between key BER proteins and cofactors [4, 9, 12,13,14,15,16,17,18]. The formation of hairpins in a TNR tract can block 5’-flap cleavage by flap endonuclease 1 (FEN1), resulting in alternate flap cleavage in which FEN1 removes fewer.
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