title: Incidence and persistence of 8-oxo-7,8-dihydroguanine within a hairpin intermediate exacerbates a toxic oxidation cycle associated with trinucleotide repeat expansion.
abstract: The repair protein 8-oxo-7,8-dihydroguanine glycosylase (OGG1) initiates base excision repair (BER) in mammalian cells by removing the oxidized base 8-oxo-7,8-dihydroguanine (8-oxoG) from DNA. Interestingly, OGG1 has been implicated in somatic expansion of the trinucleotide repeat (TNR) sequence CAG/CTG. Furthermore, a 'toxic oxidation cycle' has been proposed for age-dependent expansion in somatic cells. In this cycle, duplex TNR DNA is (1) oxidized by endogenous species; (2) BER is initiated by OGG1 and the DNA is further processed by AP endonuclease 1 (APE1); (3) a stem-loop hairpin forms during strand-displacement synthesis by polymerase β (pol β); (4) the hairpin is ligated and (5) incorporated into duplex DNA to generate an expanded CAG/CTG region. This expanded region is again subject to oxidation and the cycle continues. We reported previously that the hairpin adopted by TNR repeats contains a hot spot for oxidation. This finding prompted us to examine the possibility that the generation of a hairpin during a BER event exacerbates the toxic oxidation cycle due to accumulation of damage. Therefore, in this work we used mixed-sequence and TNR substrates containing a site-specific 8-oxoG lesion to define the kinetic parameters of human OGG1 (hOGG1) activity on duplex and hairpin substrates. We report that hOGG1 activity on TNR duplexes is indistinguishable from a mixed-sequence control. Thus, BER is initiated on TNR sequences as readily as non-repetitive DNA in order to start the toxic oxidation cycle. However, we find that for hairpin substrates hOGG1 has reduced affinity and excises 8-oxoG at a significantly slower rate as compared to duplexes. Therefore, 8-oxoG is expected to accumulate in the hairpin intermediate. This damage-containing hairpin can then be incorporated into duplex, resulting in an expanded TNR tract that now contains an oxidative lesion. Thus, the cycle restarts and the DNA can incrementally expand.
authors: Department of Chemistry, Brown University, Providence, RI 02912, USA.
authors: Daniel A
authors: DA
authors: Jarem
authors: Nicole R
authors: NR
authors: Wilson
authors: Kelly M
authors: KM
authors: Schermerhorn
authors: Sarah
authors: S
authors: Delaney
mesh_terms: D001483:Base Sequence; D045647:DNA Glycosylases / Q000737:chemistry; D057075:Enzyme Assays; D006147:Guanine / Q000031:analogs & derivatives* / Q000737:chemistry; D006801:Humans; D055029:Inverted Repeat Sequences; D007700:Kinetics; D009690:Nucleic Acid Conformation; D010084:Oxidation-Reduction; D011089:Polydeoxyribonucleotides / Q000737:chemistry; D011485:Protein Binding; D019680:Trinucleotide Repeat Expansion
chemical_list: D011089:Polydeoxyribonucleotides; C024829:8-hydroxyguanine; D006147:Guanine; D045647:DNA Glycosylases; C479058:oxoguanine glycosylase 1, human