Concentration dependent alterations of DNA replication initiation and elongation by benzo[a]pyrene diol epoxide

Vero cells treated with various concentrations of (+/-)7 beta, 8 alpha-dihydroxy-9 alpha, 10 alpha-epoxy, 7,8,9,10-dihydrobenzo[a]pyrene (BP-diol epoxide I) exhibited dose-dependent inhibition in both the rate of DNA synthesis and in the size of nascent DNA. The maximum inhibition was seen 2--3 h after addition of BP-diol epoxide I. A recovery in both the rate of synthesis and size of nascent DNA was observed 5--10 h after treatment. The pH step alkaline elution assay which separates different nascent DNA replication intermediates was used to investigate whether the inhibition and recovery noted above could be accounted for by alterations in DNA replication initiation (DNA synthesis within a replicon) or elongation (rejoining of replicons). At lowest dose studied (0.166 muM BP-diol epoxide I) a reversible inhibition in DNA initiation was observed. At the higher dose levels (0.66 muM and 1.66 muM BP-diol epoxide I) inhibition of both DNA initiation and elongation were observed and inhibition of elongation predominated. The inhibition in elongation was detected by an increase in the relative amount of low molecular weight nascent DNA associated with DNA synthesis within a replicon and a relative decrease in the higher molecular weight elongated DNA. A reversal in the inhibition of both initiation and elongation was noted.