EFFECTS OF CHEMICAL CARCINOGENS ON TRANSFORMING DNA SEQUENCES AND EXPRESSION

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The aim of this project is to identify, characterize, and clone those genes that drive the development of neoplasia whose malignant potential results from changes caused by chemical carcinogens. (A). DNAase I-hypersensitive (HS) sites were identified as targets for rapid binding and repair following in vivo exposure to benzo(a)pyrene (BP), both in total liver cell DNA and in the c-Ha-ras-1 protooncogene. The kinetics of BP adduct formation and repair were first determined in total liver DNA from hamsters given tritiated BP intraportally. Isolation of nuclei at selected times from BP treatment showed that 80% of the adducts were DNAase I-HS at early times after BP exposure (30 min), whereas the adducts remaining when repair was 90% complete (60 min) were no longer DNAase I-HS. The Ha-ras gene was analyzed under the same conditions of BP exposure in hamster liver DNA and showed a marked DNAase I-HS response at the early time points (15-30 min), but not after repair completion (120 min), indicating that BP binding and repair occur preferentially at DNAase I-HS sites. DNAase I-HS sites were also found in the Ha-ras locus in human liver DNA, where two such sites were recognized, probably in the promoter and enhancer regions. (B). The gene encoding the polypeptide that forms the brain amyloid in Alzheimer's disease and in adult Down's syndrome was isolated, characterized and sequenced. The gene was identified by synthesis of a 59-oligonucleotide probe with deoxyinosine in every third position, hybridization to a clone from a human brain cDNA library, and sequencing. This gene was found to encode a single mRNA species, to be transcribed in normal tissues, to be conserved in distant species and to be localized in chromosome 21.