SLOW REPAIR OF BULKY DNA-ADDUCTS ALONG THE NONTRANSCRIBED STRAND OF THE HUMAN P53 GENE MAY EXPLAIN THE STRAND BIAS OF TRANSVERSION MUTATIONS IN CANCERS

Using UvrABC incision in combination with ligation-mediated PCR (LMPCR ) we have previously shown that benzo(a)pyrene diol epoxide (BPDE) add uct formation along the nontranscribed strand of the human p53 gene is highly selective; the preferential binding sites coincide with the ma jor mutation hotspots found in human lung cancers. Both sequence-depen dent adduct formation and repair may contribute to these mutation hots pots in tumor tissues. To test this possibility, we have extended our previous studies by mapping the BPDE adduct distribution in the transc ribed strand of the p53 gene and quantifying the rates of repair for i ndividual damaged bases in exons 5, 7, and 8 for both DNA strands of t his gene in normal human fibroblasts. We found that: (i) on both stran ds, BPDE adducts preferentially form at CpG sequences, and (ii) repair of BPDE adducts in the transcribed DIVA strand is consistently faster than repair of adducts in the nontranscribed strand, while repair at the major damage hotspots (guanines at codons 157, 248 and 273) in the nontranscribed strand is two to four times slower than repair at othe r damage sites. These results strongly suggest that both preferential adduct formation and slow repair lead to hotspots for mutations at cod ons 157, 248 and 273, and that the strand bias of bulky adduct repair is primarily responsible for the strand bias of G to T transversion mu tations observed in the p53 gene in human cancers.