Mutation spectra of chemical mutagens determined by Lac+ reversion assay with Escherichia coli WP3101P-WP3106P tester strains.
We previously reported the development of mutation-specific Escherichia coli B tester strains WP3101 to WP3106 from strain WP2uvrA. In this study we constructed their pKM101-containing derivatives WP3101P to WP3106P, and further isolated their rfa derivatives WP4101-WP4106 and WP4101P-WP4106P. The six kinds of F' plasmids (lacI-, lacZ-, proAB+), each of which carries a different lacZ allele, contained in the above strains were originally derived from E. coli K-12 strains CC101-CC106. All the tester strains show Lac- and Trp- phenotype. Assays for transitions and transversions are based upon Lac+ reversion of a specific mutation located within the lacZ gene on an F' plasmid. The trpE65(ochre) allele in the same strains enables them to be used for Trp+ reversion assays as well. In the present paper, we evaluated the sensitivity, specificity, and usefulness of the newly developed tester strains. Strains WP3101P-WP3106P were highly sensitive to determine mutational profile of heterocyclic amines with S9 mix-mediated metabolic activation and most of the oxidative mutagens and free radical generators tested. Every type of base-pair substitutions induced by 2-amino-3,4-dimethylimidazo[4,5-f]quinoline (MeIQ) or 5-diazouracil were detected in strains WP3101P-WP3106P, while A:T-->C:G and G:C-->A:T mutations induced by MeIQ, and A:T-->C:G, G:C-->A:T, and G:C-->C:G by 5-diazouracil were not detected in pKM101-free tester strains. In pKM101-carrying strains, cumene hydroperoxide induced all types of base substitutions, while formaldehyde preferentially induced G:C-->T:A transversions. Phenazine methosulfate induced predominantly G:C-->A:T transitions and G:C-->T:A transversions, while H2O2 induced predominantly G:C-->T:A and A:T-->T:A transversions. Introduction of the rfa mutation considerably enhanced sensitivity to bulky mutagens such as polycyclic aromatic compounds. All six possible base substitutions induced by 9, 10-dimethyl-1,2-benzanthracene (DMBA) were detected in tester strains WP4101P-WP4106P. In conclusion, our tester strains WP3101P-WP3106P and WP4101P-WP4106P permitted rapid and simple detection of specific mutations induced by variety of mutagens.
Ohta
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School of Life Science, Tokyo University of Pharmacy and Life Science, 1432-1 Horinouchi, Hachioji, Tokyo 192-0392, Japan. ohta@1s.toyaku.ac.jp
Watanabe-Akanuma
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Tokishita
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Yamagata
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D000818:Animals; D002457:Cell Extracts; D004269:DNA, Bacterial / Q000235:genetics; D004353:Drug Evaluation, Preclinical / Q000379:methods; D004926:Escherichia coli / Q000187:drug effects* / Q000235:genetics / Q000254:growth & development; D016368:Frameshift Mutation; D006571:Heterocyclic Compounds / Q000633:toxicity; D007763:Lac Operon / Q000187:drug effects* / Q000235:genetics; D008099:Liver / Q000378:metabolism; D008297:Male; D016254:Mutagenesis, Insertional / Q000379:methods; D009152:Mutagenicity Tests / Q000379:methods*; D009153:Mutagens / Q000737:chemistry / Q000633:toxicity*; D011084:Polycyclic Aromatic Hydrocarbons / Q000633:toxicity; D051381:Rats; D017207:Rats, Sprague-Dawley; D012680:Sensitivity and Specificity; D015262:Xenobiotics / Q000737:chemistry / Q000633:toxicity*
D002457:Cell Extracts; D004269:DNA, Bacterial; D006571:Heterocyclic Compounds; D009153:Mutagens; D011084:Polycyclic Aromatic Hydrocarbons; D015262:Xenobiotics