Identification and characterization of FGF target genes

FGF 靶基因的鉴定和表征

• 批准号: 9556525
• 负责人: MARK B LEWANDOSKI
• 金额: $ 23.77万
• 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/9556525
• 关键词: Address; Alpha Cell; Area; Behavior; Binding; Biological Assay; Breast Cancer Genetics; Cardiac; Cell Culture System; Cell Culture Techniques; Cell Death; Cells; Chronic Kidney Failure; Cis-Acting Sequence; Complement Factor B; DNA; Data; Development; Differentiated Gene; Disease; Elements; Embryo; Embryonic Development; Fibroblast Growth Factor; Gene Expression; Gene Targeting; Genes; Genomics; Goals; In Vitro; Knowledge; Libraries; Malignant Neoplasms; Malignant neoplasm of prostate; Molecular; Mus; Oncogenes; Pathology; Play; Proteins; Reporter Genes; Research; Role; Signal Pathway; Signal Transduction; System; Techniques; Trans-Activators; Transfection; Urologic Diseases; angiogenesis; arm; base; bone; cancer site; cis acting element; drug development; experimental study; in vivo; insight; malignant breast neoplasm; migration; mouse genome; mutant; neurogenesis; novel

While we can readily identify FGF targets genes, to address the deficiency in our knowledge of what why genes respond to FGF signaling, we have initiated 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. 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 FGF signals, in the developing mouse embryo. Transfection experiments reveal that these sequences are also FGF responsive in culture cells. Now the goals are to a) determine the minimal FGF 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.

虽然我们可以很容易地确定FGF靶基因，以解决我们的知识不足，为什么基因响应FGF信号，我们已经启动了一个项目，分子定义的顺式作用元件，导致FGF靶基因响应FGF信号。该项目的一个分支涉及表征调节Sprouty2和Sprouty4表达的元件，这两个充分表征的FGF靶基因编码调节FGF信号通路的蛋白质。在我们的初步研究中，我们已经定义了来自Sprouty2的序列，这些序列驱动胚胎FGF信号传导关键区域的表达。使用项目ZIA BC 010338（脊椎动物发育中Fgf信号传导的作用）中描述的突变体，我们已经确定在发育中的小鼠胚胎中，由这些序列驱动的报告基因表达对FGF信号有响应。转染实验表明，这些序列在培养细胞中也是FGF响应性的。现在的目标是a）确定最小的FGF 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，定义细胞培养系统，在其中测定FGF反应性。这样的系统将被用来，在我们的第二个手臂的这个项目，筛选新的库，以确定区域的FGF反应在小鼠基因组。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。