HIGHER ORDER PROTEIN-DNA COMPLEXES IN GENE REGULATION

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The goal of this project is to discover how "winged helix" transcription factors reorganize and position nucleosome structure, thereby creating higher-order protein-DNA complexes that regulate gene expression. Regulatory factors can function on positioned nucleosomes at transcriptional enhancer elements, yet it is unknown how nucleosomes become positioned at the appropriate developmental time and place, and whether they are required for transcriptional activation. The investigator can now address these issues, give some recent advances: the investigator's studies discovered that binding of the winged helix factor HNF3 is sufficient to position a nucleosome core at the transcriptional enhancer of the serum albumin gene; have defined specific amino-acids within and outside the winged helix DNA binding domain that are critical for HNF3 binding to nucleosome cores and to affect particle structure and have devised systems to critically assess the relationship between nucleosome reorganizing and chromatin function in vivo. The P.I. proposes to extend these findings with the following specific aims: 1) To investigate the biochemical mechanism by which HNF3 influences the position of a nucleosome core along the DNA. 2) To reveal how winged helix proteins affect DNA access of the nucleosome core and thereby enhance or inhibit the binding of other proteins. 3) To selectively perturb the nucleosome reorganizing properties of HNF3 and assess the functional consequences in endogenous cell chromatin.