The overall aim of the research proposed here is to understand how non-B DNA structures rise at specific sites in chromosomal DNA (which is hyperactive to certain chemical carcinogens) and to understand the role of these non-B DNA structures in gene expression. Such DNA structure has been detected with a suspected carcinogen, bromoacetalehyde (BAA), which is known to react specifically with unpaired DNA bases. Recently, related compound, the known chemical carcinogen, chloroacetaldehyde (CAA), was also found to react at the same sites in chromatin as were detected with BAA. When regulatory regions of genes in supercoiled plasmid DNA are examined using BAA and CAA, multiple DNA sequences are found which, under torsional stress, readily adopt non-B structure. The DNA sequences with this physical property are operationally defined in his proposal as non-B sequences. The CAA-reactive sites in chromatin are usually found within or in close proximity to these non-B sequences. This proposal will test the hypothesis that non- B sequences play an important role in active chromatin assembly nd thus in regulation of gene expression. To do this, the unusual physical properties of non-B sequences found in the mouse immunoglobulin JH-C MuM intron will be analyzed. The mechanism by which certain non-B sequences argument level of gene expression similarly to the polyoma enhancer when placed in an expression system) will be investigated. The functional properties of certain non-B sequences and known enhancers will be compared. The possibilities for non-B sequences affecting nucleosome phasing, RNA transcription initiation sites, and the interaction of host cellular factors with their neighboring DNA sequences will be studied. The effect of DNA template topology on factor binding potential will be determined. A new minichromosome expression system to study chromatin structure-function relationships will be established. The physical and functional analysis of non-B sequences should facilitate understanding of the biological significance of the target sites for certain chemical carcinogens and thus the mechanism by which a carcinogen causes cancer.
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