Repair of oxidatively damaged guanine in Saccharomyces cerevisiae by an alternative pathway.
BACKGROUND
Transversion mutations are caused by 8-oxoguanine (OG), a DNA lesion produced by the spontaneous oxidation of guanine nucleotides, which mis-pairs with adenine during replication. Resistance to this mutagenic threat is mediated by the GO system, the components of which are functionally conserved in bacteria and mammals. To date, only one of three GO system components has been identified in the budding yeast Saccharomyces cerevisiae, namely the OG:C-specific glycosylase/lyase yOgg1. Furthermore, S. cerevisiae has been reported to contain a unique glycosylase/lyase activity, yOgg2, which excises OG residues opposite adenines. Paradoxically, according to the currently accepted model, yOgg2 activity should increase the mutagenicity of OG lesions. Here we report the isolation of yOgg2 and the elucidation of its role in oxidative mutagenesis.


RESULTS
Borohydride-dependent cross-linking using an OG-containing oligonucleotide substrate led to the isolation of yOgg1 and a second protein, Ntg1, which had previously been shown to process oxidized pyrimidines in DNA. We demonstrate that Ntg1 has OG-specific glycosylase/lyase activity indistinguishable from that of yOgg2. Targeted disruption of the NTG1 gene resulted in complete loss of yOgg2 activity and yeast lacking NTG1 had an elevated rate of A:T to C:G transversions.


CONCLUSIONS
The Ntg1 and yOgg2 activities are encoded by a single gene. We propose that yOgg2 has evolved to process OG:A mis-pairs that have arisen through mis-incorporation of 8-oxo-dGTP during replication. Thus, the GO system in S. cerevisiae is fundamentally distinct from that in bacteria and mammals.
Bruner
S D
SD
Harvard University, Department of Chemistry and Chemical Biology, 12 Oxford Street, Cambridge, Massachusetts 02138, USA.
Nash
H M
HM
Lane
W S
WS
Verdine
G L
GL
D000345:Affinity Labels; D000595:Amino Acid Sequence; D001483:Base Sequence; D001894:Borohydrides; D017931:DNA Primers / Q000235:genetics; D004260:DNA Repair / Q000235:genetics; D004271:DNA, Fungal / Q000737:chemistry / Q000378:metabolism; D043603:DNA-(Apurinic or Apyrimidinic Site) Lyase; D043382:DNA-Formamidopyrimidine Glycosylase; D018390:Gene Targeting; D005800:Genes, Fungal; D006147:Guanine / Q000031:analogs & derivatives* / Q000737:chemistry / Q000378:metabolism; D008969:Molecular Sequence Data; D016296:Mutagenesis; D009699:N-Glycosyl Hydrolases / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D010084:Oxidation-Reduction; D011994:Recombinant Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity
D000345:Affinity Labels; D001894:Borohydrides; D017931:DNA Primers; D004271:DNA, Fungal; D011994:Recombinant Proteins; D029701:Saccharomyces cerevisiae Proteins; C024829:8-hydroxyguanine; D006147:Guanine; D009699:N-Glycosyl Hydrolases; D043382:DNA-Formamidopyrimidine Glycosylase; D043603:DNA-(Apurinic or Apyrimidinic Site) Lyase; C101825:NTG1 protein, S cerevisiae