Mutagenic specificity of the food mutagen 2-amino-3-methylimidazo[4,5-f]quinoline in Escherichia coli using the yeast URA3 gene as a target.
2-Amino-3-methylimidazo[4,5-f]quinoline (IQ), a strong mutagen/carcinogen, belongs to a group of heterocyclic amines that are formed (ng/g amounts) during the cooking of protein containing food. The mutational specificity of IQ in Escherichia coli was determined in a forward mutation assay using the yeast URA3 gene as a target. The plasmid pTU-AC, containing the target URA3, was randomly modified in vitro using N-hydroxy-IQ, and subsequently transformed into an E. coli pyrF strain (DB6656). Mutant clones were directly selected by their ability to grow on medium containing 5-fluoro-orotic acid which is toxic to URA3+ clones and thereby selects for URA3- mutants. Single Strand Conformation Polymorphism (SSCP) was used to map the mutation-containing regions of URA3, so that it was necessary to sequence only the relevant, mutation-containing fragment and not the entire gene. At a modification level of 7 IQ-lesions/URA3 gene, the predominant mutations were base substitutions (approximately 70%), followed by complex gene rearrangements (approximately 20%) and frameshifts (approximately 10%). More than 96% of the base substitutions occurred at G:C base pairs and were predominantly G:C-->A:T transitions, followed by G:C-->T:A and G:C-->C:G transversions. Next neighbour analysis revealed that deoxyguanosines situated within the sequence 5'-TGC were more susceptible to mutations induced by IQ. With one exception, all frameshift mutations were -1 deletions at runs of three consecutive dGs. At higher IQ-modification levels, predominantly complex sequence rearrangements were observed.
Broschard
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TH
Unité de Canćerogenèse et Mutagenèse Moléculaire et Structuale/CNRS UPR 9003, Pôle API, Ecole Supérieure de Biotechnologie de Strasbourg, Illkirch-Graffenstaden, France.
Lebrun-Garcia
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A
Fuchs
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RP
D001483:Base Sequence; D004926:Escherichia coli / Q000187:drug effects / Q000235:genetics*; D005656:Fungal Proteins / Q000187:drug effects / Q000235:genetics*; D009152:Mutagenicity Tests; D009153:Mutagens / Q000494:pharmacology*; D018807:Polymorphism, Single-Stranded Conformational; D011804:Quinolines / Q000494:pharmacology*; D013045:Species Specificity; D015003:Yeasts / Q000235:genetics
D005656:Fungal Proteins; D009153:Mutagens; D011804:Quinolines; C029216:2-amino-3-methylimidazo(4,5-f)quinoline