Expression of a Human Cytochrome P450 in Yeast Permits Analysis of Pathways for Response to and Repair of Aflatoxin-Induced DNA Damage

Abstract

ABSTRACT Aflatoxin B 1 (AFB 1 ) is a human hepatotoxin and hepatocarcinogen produced by the mold Aspergillus flavus . In humans, AFB 1 is primarily bioactivated by cytochrome P450 1A2 (CYP1A2) and 3A4 to a genotoxic epoxide that forms N 7 -guanine DNA adducts. A series of yeast haploid mutants defective in DNA repair and cell cycle checkpoints were transformed with human CYP1A2 to investigate how these DNA adducts are repaired. Cell survival and mutagenesis following aflatoxin B 1 treatment was assayed in strains defective in nucleotide excision repair (NER) ( rad14 ), postreplication repair (PRR) ( rad6 , rad18 , mms2 , and rad5 ), homologous recombinational repair (HRR) ( rad51 and rad54 ), base excision repair (BER) ( apn1 apn2 ), nonhomologous end-joining (NHEJ) ( yku70 ), mismatch repair (MMR) ( pms1 ), translesion synthesis (TLS) ( rev3 ), and checkpoints ( mec1 - 1 , mec1-1 rad53 , rad9 , and rad17 ). Together our data suggest the involvement of homologous recombination and nucleotide excision repair, postreplication repair, and checkpoints in the repair and/or tolerance of AFB 1 -induced DNA damage in the yeast model. Rev3 appears to mediate AFB 1 -induced mutagenesis when error-free pathways are compromised. The results further suggest unique roles for Rad5 and abasic endonuclease-dependent DNA intermediates in regulating AFB 1 -induced mutagenicity.