There is compelling evidence that growth factors regulate both cell proliferation and differentiation, but little is known about the mechanisms that regulate growth factor production. The fibroblast growth factor k-FGF is one of the first growth factors to be produced during mammalian development and it is produced by a wide range of tumors. Recently, embryonal carcinoma (EC) cells have been used to study the expression and regulation of the k-FGF oncogene. EC cells, which serve as an excellent model system for studying early development, have been shown to produce k-FGF; whereas their differentiated cells do not. Efforts to understand how k-FGF expression is regulated indicate that differentiation represses transcription of this gene. This conclusion is supported by nuclear run-off studies and by the differential expression of promoter/reporter gene constructs in EC cells and their differentiated cells. Based on the most recent studies in the PI's laboratory, it is hypothesized that expression of the k-FGF gene in EC cells and their differentiated cells is controlled by at least three cis-regulatory elements: an essential enhancer in the third exon of the gene, a positive cis-regulatory element and a negative cis-regulatory element located upstream of the transcription start site. To test this hypothesis and to determine how expression of this gene is regulated, four Specific Aims are proposed: 1) Identify and map precisely the location of cis-regulatory elements that control expression of the k-FGF gene in mouse EC cells and their differentiated cells. 2) Examine the binding of nuclear proteins to the cis-regulatory elements that control expression of the k-FGF gene. 3) Identify the transcription factors that regulate expression of this gene. 4) Determine how differentiation of EC cells regulates expression of the transcription factors that regulate expression of the k-FGF gene. The work proposed in this application will address a critical gap in our current knowledge, namely, our limited understanding of the mechanisms by which growth factor production is regulated. Equally important, this work will help advance our understanding of how differentiation regulates the expression of transcription factors.
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