REGULATION OF GROWTH FACTORS AND MAMMALIAN EMBRYOGENESIS

有令人信服的证据表明，生长因子调节这两种细胞 增殖和分化，但对其了解甚少 调节生长因子生产的机制。成纤维细胞的生长 因子k-成纤维细胞生长因子是第一批在 哺乳动物的发育和它是由多种肿瘤产生的。 近年来，胚胎癌(EC)细胞已被用于研究其生物学行为。 K-成纤维细胞生长因子癌基因的表达和调控。EC细胞，它服务于 作为研究早期发展的优秀模型系统，已经被 它们的分化细胞不产生k-成纤维细胞生长因子。 了解k-成纤维细胞生长因子表达是如何调控的努力表明 分化抑制了该基因的转录。这个结论是 由核径流研究和差异表达支持 启动子/报告基因在EC细胞中的表达及其分化 细胞。根据PI实验室的最新研究，它是 推测k-成纤维细胞生长因子基因在EC细胞中的表达及其表达 分化的细胞受至少三种顺式调控因子的控制 元素：基因第三外显子的基本增强子，阳性 顺式调控元件和负顺调控元件位于 转录起始点的上游。来检验这一假设，并 确定该基因的表达是如何调节的，有四个具体目标 建议：1)准确识别和绘制 调控人成纤维细胞生长因子基因表达的顺式调控元件 小鼠EC细胞及其分化的细胞。2)检查 核蛋白与控制表达的顺式调控元件 K-fgf基因。3)确定调控转录因子 这种基因的表达。4)确定EC细胞如何分化 调节转录因子的表达，从而调节 K-成纤维细胞生长因子基因表达。本申请中提出的工作将 解决我们当前知识中的一个关键差距，即我们有限的 对生长因子产生的机制的理解 受监管的。同样重要的是，这项工作将有助于推进我们的 了解分化是如何调控基因表达的 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.