Formation and Repair of Mismatches Containing Ribonucleotides and Oxidized Bases at Repeated DNA Sequences.
The cellular pool of ribonucleotide triphosphates (rNTPs) is higher than that of deoxyribonucleotide triphosphates. To ensure genome stability, DNA polymerases must discriminate against rNTPs and incorporated ribonucleotides must be removed by ribonucleotide excision repair (RER). We investigated DNA polymerase β (POL β) capacity to incorporate ribonucleotides into trinucleotide repeated DNA sequences and the efficiency of base excision repair (BER) and RER enzymes (OGG1, MUTYH, and RNase H2) when presented with an incorrect sugar and an oxidized base. POL β incorporated rAMP and rCMP opposite 7,8-dihydro-8-oxoguanine (8-oxodG) and extended both mispairs. In addition, POL β was able to insert and elongate an oxidized rGMP when paired with dA. We show that RNase H2 always preserves the capacity to remove a single ribonucleotide when paired to an oxidized base or to incise an oxidized ribonucleotide in a DNA duplex. In contrast, BER activity is affected by the presence of a ribonucleotide opposite an 8-oxodG. In particular, MUTYH activity on 8-oxodG:rA mispairs is fully inhibited, although its binding capacity is retained. This results in the reduction of RNase H2 incision capability of this substrate. Thus complex mispairs formed by an oxidized base and a ribonucleotide can compromise BER and RER in repeated sequences.
From the Department of Environment and Primary Prevention, Istituto Superiore di Sanità, 00161 Roma and the Department of Science, University Roma Tre, 00154 Roma, Italy.
Piera
P
Cilli
From the Department of Environment and Primary Prevention, Istituto Superiore di Sanità, 00161 Roma and.
Anna
A
Minoprio
From the Department of Environment and Primary Prevention, Istituto Superiore di Sanità, 00161 Roma and.
Cecilia
C
Bossa
From the Department of Environment and Primary Prevention, Istituto Superiore di Sanità, 00161 Roma and margherita.bignami@gmail.com.
Margherita
M
Bignami
From the Department of Environment and Primary Prevention, Istituto Superiore di Sanità, 00161 Roma and.
Filomena
F
Mazzei
D020137:Base Pair Mismatch; D004247:DNA / Q000737:chemistry* / Q000378:metabolism; D019951:DNA Polymerase beta / Q000378:metabolism; D010084:Oxidation-Reduction; D012265:Ribonucleotides / Q000737:chemistry* / Q000378:metabolism
D012265:Ribonucleotides; D004247:DNA; D019951:DNA Polymerase beta
8-oxoguanine (8-oxoG); 8-oxoguanine glycosylase (OGG1); DNA damage; DNA polymerase; DNA repair; mutY homolog (MUTYH); ribonuclease