Carcinogenesis, Vol 19, 457-461, Copyright © 1998 by Oxford University Press
GT Pauly, SH Hughes and RC Moschel
To compare mutagenesis by O6-methylguanine (m6G), O4-methylthymine (m4T)
and O6-ethylguanine (e6G), and assess their genotoxicity in Escherichia
coli, double-stranded and gapped plasmids were constructed containing a
single m6G, e6G or m4T in the initiation codon (ATG) of a lacZ' gene.
Modified base induced mutations were scored by the loss of lacZ' activity
on X-gal-containing media resulting in formation of white or sectored
(mutant) rather than blue (non-mutant) colonies. Genotoxicity experiments
with gapped plasmids containing the modified bases indicated that m4T
produced a greater number of bacterial colonies than m6G or e6G. m4T was
more mutagenic (45% mutant colonies) than m6G (6%) or e6G (11%) in repair
competent (w.t.) E. coli when incorporated in double-stranded plasmids. In
gapped plasmids, m4T produced 99% mutant colonies (as was observed
previously for e6G) in both w.t. E. coli or E. coli deficient in both
O6-alkylguanine-DNA alkyltransferases as well as methylation-directed
mismatch repair (ada(- )-ogt(-)-mutS[-]). m6G in gapped plasmids produced
62% mutant colonies in w.t. E. coli, but this percentage increased to 94%
in the ada(-)- ogt(-)-mutS(-) strain. In double-stranded plasmids both m4T
and m6G produced very similar distributions of mutant and non-mutant
colonies in the ada(-)-ogt(-)-mutS(-) strain. These observations led to the
conclusion that differences in the mutagenicity of m6G and m4T in w.t. E.
coli were a result of preferential repair of m6G compared to m4T by
alkyltransferase and mismatch repair mechanisms, and did not reflect
differences in their respective coding efficiency or their inherent
obstructiveness to DNA synthesis as was observed with e6G. The combination
of alkyltransferase and mismatch repair was concluded to be primarily
responsible for the apparent genotoxicity of m6G compared to m4T in
double-stranded plasmids.
ARTICLES
Comparison of mutagenesis by O6-methyl- and O6-ethylguanine and O4- methylthymine in Escherichia coli using double-stranded and gapped plasmids
Chemistry of Carcinogenesis Laboratory, ABL-Basic Research Program, NCI- Frederick Cancer Research and Development Center, MD 21702, USA.
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