Environmental carcinogens of the polycyclic aromatic hydrocarbon class, such as benzo(a)pyrene, are metabolized by cellular enzyme systems to highly reactive bay-region diol epoxides which have been identified as ultimate carcinogens. These compounds modify macromolecules in cells, thereby initiating the process of carcinogenesis. Although the interactions of benzo(a)pyrene diol epoxide (BPDE-I) with DNA have been studied in vitro, little is known of how this relates to interactions which take place in cellular nuclei, where the DNA is intimately associated with a variety of chromosomal proteins. Even less is known of the interactions of BPDE-I with these proteins, nor of how such interactions may affect the regulation of gene expression which involves interactions between DNA control regions and trans-acting proteins. In the present proposal, several model systems will be studied in which a particular DNA-binding protein and its target DNA can be prepared in large amounts utilizing recombinant DNA technology. The systems include the interaction of histone proteins with DNA, the interaction of the human transcription factor Sp1 with the promoter of the aprt gene, the interaction of yeast transaction factor SW15 with its target DNA in the HO gene, and the interaction of the Xenopus transcription factor TFIIIA with the internal control region of the 5S RNA gene. The latter three are all examples of zinc-finger transcription factors, which appear to be of widespread importance. The following questions will be asked of these systems: 1) In nucleosomal complexes of the histones with DNA, where are DNA binding sites for BPDE-I located? 2) How does binding of the protein to its target DNA affect the formation of covalent BPDE-I DNA adducts? 3) Does BPDE-I bind covalently to the transcription factors? If so, what are the target amino acid residues and how is this binding affected by interaction with the target DNA sequence? 4) What is the effect of BPDE-I-DNA adduct formation in a target sequence upon the subsequent interaction with its cognate transcription factor?
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