Extrachromosomal probes for mutagenesis by carcinogens: studies on the mutagenic activity of O6-methylguanine built into a unique site in a viral genome.
This work examines the mutagenic activity of O6-methylguanine (O6MeGua), a DNA adduct formed by certain carcinogenic alkylating agents. A tetranucleotide, 5'-HOTpm6GpCpA-3', was synthesized and ligated into a four-base gap in the unique Pst I site of the duplex genome of the E. coli virus, M13mp8. The double-stranded ligation product was converted to single-stranded form and used to transform E. coli to produce progeny phage. The mutation frequency of O6MeGua was defined as the percentage of progeny phage with mutations in their Pst I site, and this value was determined to be 0.4%. To determine the impact of DNA repair on mutagenesis, cellular levels of O6MeGua-DNA methyltransferase (an O6MeGua-repair protein) were depleted by treatment of host cells for virus replication with N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) prior to viral DNA uptake. In these host cells, the mutation frequency due to O6MeGua increased markedly with increasing MNNG dose (the highest mutation frequency observed was 20%). DNA sequence analysis of mutant genomes revealed that in both MNNG treated and untreated cells, O6MeGua induced exclusively G to A transitions.
D001483:Base Sequence; D002273:Carcinogens / Q000633:toxicity*; D003090:Coliphages / Q000187:drug effects / Q000235:genetics*; D004260:DNA Repair; D004926:Escherichia coli / Q000187:drug effects / Q000235:genetics*; D005814:Genes, Viral / Q000187:drug effects*; D006147:Guanine / Q000031:analogs & derivatives* / Q000032:analysis / Q000633:toxicity; D009153:Mutagens; D009154:Mutation; D009838:Oligodeoxyribonucleotides / Q000138:chemical synthesis
D002273:Carcinogens; D009153:Mutagens; D009838:Oligodeoxyribonucleotides; D006147:Guanine; C008449:O-(6)-methylguanine