This research program seeks to devise new strategies to rationally inhibit gene expression mediated by transcription factors (TFs). The strategy is to design modular polyamide DNA minor binding agents that potently and specifically inhibit TF binding to gene promoters and disrupt gene expression. For this purpose, a representative of the Ets oncogenic protein family Elk-1, which binds a serum response element (SRE) contained within the c-fos promoter, was chosen as a model TF. Interference with the binding of Elk-1 to the SRE will be quantitatively measured by mobility shift assays, to assess a polyamide's ability to inhibit TF/DNA complexes. Quantitative footprint analysis will be used to determine where polyamides bind on the SRE promoter element and also the strength of the binding. Polyamides that are potent and specific inhibitors of Elk-1/SRE complex formation will be evaluated for their ability to inhibit transcriptional activation from the c-fos promoter in cell-free and cellular assays. Promising agents will be evaluated for their ability to interfere with a neoplastic phenotype associated with c-fos gene expression. In parallel, polyamides will be optimized for cellular activity while maintaining target specificity.
Specific aims :
Aim1. Identification of polyamide DNA minor groove binding agents that interfere with the binding of Elk-1 to the c-fos promoter under cell-free conditions.
Aim2. Effects of polyamide DNA minor groove binding agents on Elk-1 mediated gene expression of the c-fos promoter under cell-free conditions.
Aim3. Effects of polyamide DNA minor groove binding agents on Elk-1 mediated gene expression of the c-fos promoter in intact cells.
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