Mutational spectrum of N-hydroxy-N-acetyl-4-aminobiphenyl at exon 3 of theHPRT gene

4-Aminobiphenyl is a human bladder carcinogen present in many environmental sources, including cigarette smoke. It can be metabolized in two steps to the mutagen N-hydroxy-N-acetyl-4-aminobiphenyl (N-OH-AABP). In this study t he mutational spectrum of N-OH-AABP-exposed human lymphoblastoid cells (TK6 ) was determined using HPRT as the target gene. Three large, HAT (hypoxanth ine-aminopterin-thymidine)-cleaned TK6 cultures were independently treated with 20 muM N-OH-AABP for 24 h, allowed to recover for 4 days, then continu ously exposed to 40 muM 6-thioguanine to select for induced mutants. Contem porary control cultures received vehicle in place of N-OH-AABP. N-OH-AABP t reatment gave an 11-fold increase in mutation frequency. Mutations were del ineated in exon 3 of the HPRT gene directly from genomic DNA extracted from both treated and untreated cells using the polymerase chain reaction, dena turing gradient gel electrophoresis (DGGE) and dideoxy sequencing. DGGE ana lysis showed N-OH-AABP increased both the number and type of mutations as c ompared with controls. The major background mutation was a G(197) --> A tra nsition. The major N-OH-AABP-induced changes were G(209-211) --> T transver sions (30 %), a seven-base repeat at position 185 (17 %) and an A(215) --> T transversion (2 %). The shift in the control spectrum of a transition to that of transversions and insertions suggest that the electrophile N-OH-AAB P forms bulky adducts at the same sites on exon 3 of HPRT as do many other bulky electrophiles, causes replication errors by similar mechanisms, but i nduces at least one potentially signature mutation.