REGULATION OF GROWTH FACTORS AND MAMMALIAN EMBRYOGENESIS

有令人信服的证据表明，生长因子调节两种细胞 增殖和分化，但很少有人知道， 调节生长因子产生的机制。 成纤维细胞生长 因子k-FGF是在生长期间产生的第一种生长因子之一。 哺乳动物的发育，它是由广泛的肿瘤。 最近，胚胎癌（EC）细胞已被用于研究肿瘤的生长。 K-FGF癌基因的表达和调节。 EC细胞， 作为研究早期发育的一个优秀模型系统， 显示产生k-FGF;而它们的分化细胞不产生。 努力了解k-FGF表达是如何调节的表明， 分化抑制该基因的转录。 这个结论是 支持核径流研究和差异表达 启动子/报告基因构建体在EC细胞中的表达及其分化 细胞 根据PI实验室的最新研究， 假设k-FGF基因在EC细胞中表达及其 分化的细胞由至少三个顺式调节因子控制， 元件：基因的第三外显子中的必需增强子， 顺式调节元件和负顺式调节元件位于 转录起始位点的上游。 为了验证这一假设， 确定该基因的表达是如何调节的，四个具体目标 建议：1）确定并精确绘制 顺式调节元件，其控制k-FGF基因的表达， 小鼠EC细胞及其分化细胞。2)检查的约束力 控制表达的顺式调节元件 k-FGF基因。3)识别调节转录因子 这个基因的表达。4)确定EC细胞如何分化 调节转录因子的表达， k-FGF基因的表达。 本申请中提出的工作将 解决我们现有知识中的一个关键差距，即我们有限的 了解生长因子生产的机制， 监管. 同样重要的是，这项工作将有助于推动我们的 了解分化如何调节 转录因子

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.