In normal cells, the telomerase gene is not expressed and therefore telomerase activity is not present. Telomerase expression is controlled by an upstream regulator element. Sierra Sciences Inc. (SSI) scientists identified a regulatory region (promoter) upstream of the telomerase gene. This promoter region controls the expression of the telomerase gene in normal (non cancer) cells. Fine mapping of this upstream region revealed a minimal promoter containing a control element (Site C), which was responsible for repression of expression when placed upstream of a reporter gene. Cell transfection assays demonstrated that a one base pair change in Site C abrogated repression. Subsequent electrophoretic shift assays (EMSAs) demonstrated that a protein complex forms on Site C using nuclear extracts prepared from normal and cancer cells. The hypotheses for this proposal is that Site C is the major regulatory region for the telomerase gene and this element is recognized by an endogenous protein that represses the expression of telomerase in normal cells. Our goal for this Phase I proposal is to isolate and identify the repressor protein. Our long-term goal is to develop small molecule drugs to prevent the repressor from binding to the Site C target. This controlled expression will then alleviate the effects due to telomerase repression.
Thesaurus Terms: gene induction /repression, genetic regulatory element, protein purification, protein quantitation /detection, protein structure function, technology /technique development, telomerase DNA primer, binding protein, protein sequence, proteomics gel mobility shift assay, molecular cloning
