The hydantoin lesions formed from oxidation of 7,8-dihydro-8-oxoguanine are potent sources of replication errors in vivo.
Single-stranded DNA genomes have been constructed that site-specifically contain the 7,8-dihydro-8-oxo-2'-deoxyguanine (8-oxoG) oxidation products guanidinohydantoin (Gh) and the two stable stereoisomers of spiroiminodihydantoin (Sp1 and Sp2). The circular viral genomes were transfected into wild-type AB1157 Escherichia coli, and the efficiency of lesion bypass by DNA polymerase(s) was assessed. Viral progeny were analyzed for mutation frequency and type using the recently developed restriction endonuclease and postlabeling (REAP) assay. Gh was bypassed nearly as efficiently as the parent 8-oxoG but was highly mutagenic, causing almost exclusive G --> C transversions. The stereoisomers Sp1 and Sp2 were, in comparison, much stronger blocks to DNA polymerase extension and caused a mixture of G --> T and G --> C transversions. The ratio of G --> T to G --> C mutations for each Sp lesion was dependent on the stereochemical configuration of the base. All observed mutation frequencies were at least an order of magnitude higher than those caused by 8-oxoG. Were these lesions to be formed in vivo, our data show that they are absolutely miscoding and may be refractory to repair after translesion synthesis.
Henderson
Paul T
PT
Biological Engineering Division and Department of Chemistry, Massachusetts Institute of Technology, Cambridge, Massachusetts 02139, USA.
Delaney
James C
JC
Muller
James G
JG
Neeley
William L
WL
Tannenbaum
Steven R
SR
Burrows
Cynthia J
CJ
Essigmann
John M
JM
D004247:DNA / Q000138:chemical synthesis* / Q000737:chemistry; D004249:DNA Damage; D004261:DNA Replication; D004270:DNA, Circular / Q000737:chemistry; D004277:DNA, Single-Stranded / Q000737:chemistry*; D004259:DNA-Directed DNA Polymerase / Q000378:metabolism; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics; D016679:Genome, Viral; D006146:Guanidines / Q000737:chemistry*; D006147:Guanine / Q000031:analogs & derivatives* / Q000737:chemistry*; D006151:Guanosine / Q000031:analogs & derivatives* / Q000737:chemistry*; D008958:Models, Molecular; D016296:Mutagenesis; D010084:Oxidation-Reduction; D013141:Spiro Compounds / Q000737:chemistry*; D013237:Stereoisomerism; D014162:Transfection; D014779:Virus Replication
C453560:7,8-dihydro-8-oxoguanine; D004270:DNA, Circular; D004277:DNA, Single-Stranded; D006146:Guanidines; D013141:Spiro Compounds; C417602:guanidinohydantion; C408822:spiroiminodihydantoin; D006151:Guanosine; C024829:8-hydroxyguanine; D006147:Guanine; D004247:DNA; D004259:DNA-Directed DNA Polymerase