CELL-SPECIFIC HORMONAL REGULATION OF GENE EXPRESSION

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Differential gene expression is the underlying determinant for cellular diversity and differentiated cell function in complex organisms. Thus, elucidation of the basis for cell-specific gene expression is required for a molecular understanding of development and differentiation. The activation and/or repression of specific genes is triggered by external messages that act through signal transduction pathways that are largely ubiquitous. An additional level of complexity exists for genes that are regulated by hormones and growth factors; those genes can exhibit cell- specificity for induced expression as well as for basal expression. The long term goal of our research is to understand how hormonal, developmental, and tissue-specific cues integrate to precisely control the activities of complex genes. The focus of the research in this proposal is to study cell-specific hormonal induction by identifying the cell-specific elements in the chicken ovalbumin gene. This expression of this gene is restricted to the tubular gland cells of the avian oviduct and is regulated by four classes of steroid hormones and by the peptide hormone insulin. The specific hypothesis being tested is that the cell-specific expression of the ovalbumin gene is achieved via positive activation by hormones in oviduct and direct repression by unknown factors in nonoviduct tissues. The Specific Aims are to I). identify and characterize the positive cell-specific regulatory elements in the ovalbumin gene, II). clone the proteins that bind to the positive cell-specific elements, and III). identify the elements in the ovalbumin gene that repress its expression in inappropriate tissues. The identification and characterization of the relevant DNA elements will be accomplished by combination of in vivo footprinting, gel mobility shift assays, and gene transfections. The cloning of the associated proteins will be attempted by a one-hybrid yeast genetic selection system or by other techniques that rely on the DNA binding properties of the proteins. Elucidation of the underlying mechanisms involved in the generation and maintenance of differentiated cells may provide the basis for understanding how events go awry in some diseases such as cancer that involve extensive alterations in cellular phenotype and proliferation. Furthermore, the avian oviduct has the potential for producing large amounts of easily purified proteins if expression can be targeted to that organ. This has implications for research and for therapeutic and commercial purposes.