The proposed research aims to develop a chemically-based system for inducing the intracellular association of target proteins in specific cells by using nontoxic, cell permeable, synthetic ligands. Organic compounds will be chemically synthesized that induce either the heterodimerization or homodimerization of intracellular proteins involved in signal transduction, transcription, protein degradation, and programmed cell death. Experiments are planned to optimize specificity and potency of the organic compounds by synthesizing sterically demanding substituents on the dimerizing ligands and introducing compensating mutations on the dimerization domains. The cell permeable synthetic reagents will be used to: (l) activate an artificial signaling pathway coupled to an inducible gene or antisense gene, (2) activate direct transcription of a target gene, (3) induce the ubiquitin-dependent intracellular proteolysis of a target protein, (4) induce the membrane translocation of raf, a component of the mitogenic signaling pathway, and (5) induce the apoptotic death of target brain cells and thereby determine their role in brain development and function. Regulated protein dimerization with cell permeable, synthetic ligands will provide a new level of control of many fundamental processes in experimental biology. Orally active dimerizers may also function as universal drugs when used to turn on or off the synthesis of therapeutic proteins (e.g., insulin for diabetes, erythropoietin for anemia, bone morphogenetic factors for osteoporis) or therapeutic nucleic acids (e.g., antisense agents or ribozymes) in gene therapy patients.
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