Vinyl chloride is a known human and rodent carcinogen that forms several cyclic base derivatives in DNA. The mutagenic potential of these derivatives has been examined in vitro but not in vivo. One of these derivatives, N2,3-ethenoguanine (εG), is known to base pair with both cytosine and thymine during in vitro DNA synthesis, which would result in G → A transitions. To determine the base pairing specificity of this labile guanine derivative in Escherichia coli, we have developed a genetic reversion assay for guanine derivatives. The assay utilizes DNA polymerase-mediated analogue insertion into a bacteriophage vector, M13G*1, which detects all single-base substitutions at position 141 of the lacZa gene by change in plaque color. After the insertion of a single eG opposite the template cytosine at position 141 by use of edGTP and DNA polymerase and further extension with all four normal dNTPs, the DNA was transfected into E. coli. Transfection of M13G*1 containing εG at the target site yielded 135 mutants among 26,500 plaques, 134 of which represented G→ A transitions. The unconnected mutation frequency was 0.5%, as compared with the control value, ≈0.02%; when corrected for εG content and penetrance, the calculated mutagenic potential of ε(mutations/analogue) was about 13%. We thus conclude that εG specifically induces G → A transitions during DNA replication in E. coli. The M13G*1 assay may permit the testing of other labile guanine derivatives not otherwise amenable to mutagenesis studies.
|Original language||English (US)|
|Number of pages||5|
|Journal||Proceedings of the National Academy of Sciences of the United States of America|
|State||Published - 1991|
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