Abstract:
:2-Amino-3-methylimidazo[4,5-f]quinoline (IQ) and 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (MelQx) are carcinogens found in cooked meats that form DNA adducts upon metabolic activation. Purified DNA from Chinese hamster ovary (CHO) cells was reacted in vitro with the active metabolites N-acetoxy-IQ or N-acetoxy-MelQx, and the adduct levels in the 5' dihydrofolate reductase (DHFR) gene and downstream region were quantitated by Southern hybridization. Adducted and restricted DNA was treated with Escherichia coli uvrABC excinuclease or alkali (0.1 N NaOH, 37 degrees C, 60 min) to incise DNA at IQ and MelQx adduct sites. The DNA was then denatured with formamide, electrophoresed on a neutral agarose gel, transferred to a support membrane, and hybridized with sequence-specific DNA probes. Both uvrABC and alkali reduced the intensity of Southern hybridization in proportion to the number of IQ or MelQx adducts in DNA, indicating that these adducts are substrates for uvrABC and that they form alkali-labile lesions in DNA. IQ and MelQx adduct levels were the same in the 5' DHFR gene and in the downstream region. Southern hybridization analysis of pBR322 containing known levels of IQ or MelQx adducts showed that the efficiency of cutting IQ or MelQx adducts by uvrABC excinuclease and alkali was approximately 30% and 15%, respectively. 32P-postlabeling studies examining adduct level in bulk DNA further showed that the adduct profiles were identical in pBR322, CHO DNA, and cultured CHO cells exposed to the reactive metabolites of IQ or MelQx. The results indicate that IQ and MelQx adducts can be quantitated in specific genomic sequences and that this method should be directly applicable to studies of gene-specific repair of these adducts in cultured cells.
journal_name
Mol Carcinogjournal_title
Molecular carcinogenesisauthors
Nouso K,Bohr VA,Schut HA,Snyderwine EGdoi
10.1002/mc.2940070211subject
Has Abstractpub_date
1993-01-01 00:00:00pages
126-34issue
2eissn
0899-1987issn
1098-2744journal_volume
7pub_type
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