Identification and characterization of FGF target genes

FGF 靶基因的鉴定和表征

• 批准号: 8349469
• 负责人: MARK B LEWANDOSKI
• 金额: $ 20.15万
• 依托单位: DIVISION OF BASIC SCIENCES - NCI
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/8349469
• 关键词: Address; Behavior; Binding; Biological Assay; Biology; Cell Culture System; Complex; Cultured Cells; DNA; Data; Development; Disease; Elements; Embryo; Embryonic Development; Family; Fibroblast Growth Factor; Gene Expression; Gene Targeting; Genes; Goals; Human; In Vitro; Indium; Knowledge; Libraries; Limb Development; Malignant Neoplasms; Malignant neoplasm of prostate; Molecular; Pathway interactions; Play; Proteins; Reporter Genes; Role; Signal Pathway; Signal Transduction; System; Techniques; Transfection; Work; arm; base; cancer therapy; cis acting element; genetic analysis; in vivo; insight; malignant breast neoplasm; mouse genome; mutant; novel; research study; tumorigenesis

To address the deficiency in our knowledge of what defines an FGF target gene, we are initiating a project to molecularly define the cis-acting elements that cause FGF targets genes to respond to FGF signals. One arm of this project involves characterizing the elements that regulate the expression of Sprouty2 and Sprouty4, two well characterized FGF target genes that encode proteins that regulate the FGF signaling pathway. We have determined that these Sprouty genes also respond to BMP signaling in our Bmpr mutants that are described in project ZIA BC 010518 (Role of BMP and FGF signaling during limb development). Therefore we are also exploring the idea that the key control elements in Sprouty genes that may be regulated by an interaction between BMP and FGF signaling. Also , this work grows organically out of insights gained from project ZIA BC 010518. In our preliminary studies we have defined sequences derived from Sprouty2 that drive expression in key regions of embryonic FGF signaling. Using mutants described in project ZIA BC 010338 (The Role of Fgf Signaling in Vertebrate Development ) we have determined that reporter gene expression, driven by these sequences, is responsive to in vivo FGF signals. Transfection experiments reveal that these sequences are also FGF responsive in culture cells. Now the goals are to a) determine the minimal FGF and BMP responsive elements b) determine the trans-activing factors that bind to these sequences, using DNA-based techniques. We will also use these elements, as positive controls, define a cell culture system in which to assay FGF responsiveness. Such a system will be used, in our second arm of this project, to screen novel libraries to define regions of FGF responsiveness in the mouse genome. We will always validate any in vitro insights with in vivo experiments, thus providing a powerful approach that should guarantee the significance of our data.

为了解决我们对 FGF 靶基因定义知识的缺陷，我们正在启动一个项目，以分子方式定义导致 FGF 靶基因对 FGF 信号做出反应的顺式作用元件。该项目的一个部分涉及表征调节 Sprouty2 和 Sprouty4 表达的元件，这两个特征已明确的 FGF 靶基因，编码调节 FGF 信号通路的蛋白质。我们已经确定，这些 Sprouty 基因也对 Bmpr 突变体中的 BMP 信号作出反应，该突变体在项目 ZIA BC 010518（BMP 和 FGF 信号在肢体发育过程中的作用）中进行了描述。因此，我们也在探索 Sprouty 基因中的关键控制元件可能受到 BMP 和 FGF 信号传导之间相互作用的调节。此外，这项工作是从 ZIA BC 010518 项目中获得的见解有机发展出来的。在我们的初步研究中，我们定义了源自 Sprouty2 的序列，这些序列驱动胚胎 FGF 信号传导关键区域的表达。使用 ZIA BC 010338 项目（Fgf 信号在脊椎动物发育中的作用）中描述的突变体，我们确定由这些序列驱动的报告基因表达对体内 FGF 信号有反应。转染实验表明这些序列在培养细胞中也具有 FGF 反应性。现在的目标是 a) 确定最小的 FGF 和 BMP 响应元件 b) 使用基于 DNA 的技术确定与这些序列结合的反式激活因子。我们还将使用这些元素作为阳性对照，定义用于测定 FGF 反应性的细胞培养系统。在该项目的第二部分中，我们将使用这样的系统来筛选新的文库，以定义小鼠基因组中 FGF 反应性的区域。我们将始终通过体内实验来验证任何体外见解，从而提供一种强大的方法来保证我们数据的重要性。