FIBRONECTIN REGULATION AND FUNCTION IN TRANSGENIC ANIMAL

转基因动物中纤连蛋白的调控和功能

• 批准号: 2183870
• 负责人: PAMELA A NORTON
• 金额: $ 11.85万
• 依托单位: THOMAS JEFFERSON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1991
• 资助国家: 美国
• 起止时间: 1991-07-01 至 1996-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2183870
• 关键词: Escherichia coli; albumins; biological models; complementary DNA; embryo /fetus cell /tissue; extracellular matrix; fibronectins; gene expression; genetic enhancer element; genetic markers; genetic promoter element; genetic regulatory element; genetically modified animals; glycoproteins; in situ hybridization; laboratory mouse; liver; liver cells; microinjections; nucleic acid probes; plasma; polymerase chain reaction; protein sequence; protein structure function; site directed mutagenesis; transfection; transforming growth factors; western blottings

The overall goals of the proposed project are first, to develop a detailed characterization of the regulation of fibronectin gene expression and second, to identify the functions specific to the various forms of fibronectin. Fibronectins are a family of adhesive glycoproteins found in the extracellular matrix as well as in plasma. Several different protein variants are produced from a single gene by alternative splicing. Fibronectins are involved in many morphogenetic events during embryonic development, as well as in tissue remodelling events such as wound healing and inflammation-induced fibrosis. The levels and types of fibronectin synthesized vary during these processes in a cell type specific fashion. Understanding the mechanisms by which fibronectin expression is regulated thus complements studies of fibronectin functions. The liver will serve as an ideal model tissue on which to focus for the pursuit of these aims. Hepatocyte-derived fibronectin is a significant component of plasma, and this material can be deposited in a variety of other tissues as well be incorporated into blood clots. Numerous studies have reported changes in fibronectin synthesis in the liver relating to hepatic fibrosis, liver regeneration and malignancy. These goals will be achieved through the production of transgenic mice. The basis for the cell type specific regulation of fibronectin will be evaluated using a marker gene, lacZ, under the control of fragments derived from the cloned rat fibronectin gene. The pattern of expression of this easily assayable indicator gene will be interpreted in light of detailed in situ hybridization studies which will be undertaken in parallel. Particular emphasis will be placed on embryonic liver development. Through analysis of a number of individual trangenics, a picture will emerge regarding the structure of the regulatory regions of the fibronectin gene. Transgenic mice will also provide the means for achieving the other goals related to determining the functions of the various forms of fibronectin. cDNAs encoding specific fibronectin isoforms will be placed under the control of tissue specific promoters; initial experiments will target gene expression to the liver. Lines of mice will be generated which express individual forms, with the expectation that synthesis of inappropriate amounts or types of fibronectin will reveal information regarding their specific functions. More detailed studies involving mutagenesis of specific portions of FN will follow from the initial studies of two FN isoforms.

拟议项目的总体目标是首先开发详细的 纤连蛋白基因表达和调节的表征 第二，确定特定形式的功能 纤连蛋白。 纤连蛋白是在 细胞外基质以及等离子体中。 几种不同的蛋白质 变体是通过替代剪接从单个基因产生的。 纤连蛋白在胚胎中参与许多形态发生事件 开发以及组织重塑事件（例如伤口愈合） 和炎症引起的纤维化。 纤连蛋白的水平和类型 在这些过程中以特定于细胞类型的方式进行合成在这些过程中有所不同。 了解调节纤连蛋白表达的机制 因此，补充了纤连蛋白功能的研究。 肝脏将用作理想的模型组织 追求这些目标。 肝细胞衍生的纤连蛋白是重要的 血浆的组成部分，该材料可以沉积在多种 其他组织也将其掺入血凝块中。 许多研究 已经报道了与 肝纤维化，肝脏再生和恶性肿瘤。 这些目标将通过生产转基因小鼠实现。 细胞类型特异性调节纤连蛋白的基础将是 在衍生的碎片控制下使用标记基因Lacz进行评估 来自克隆的大鼠纤连蛋白基因。 表达方式 易于测定的指标基因将根据详细的解释 将并行进行的原位杂交研究。 特别重点将放在胚胎肝发育上。 通过 分析许多单独的旋齿，将出现图片 关于纤连蛋白基因的调节区域的结构。 转基因小鼠还将为实现其他目标提供手段 与确定各种形式的纤连蛋白的功能有关。 编码特定纤连蛋白同工型的cDNA将放置在 控制特定启动子；最初的实验将靶向基因 表达肝脏。 将生成鼠标线的线条 个人形式，期望综合不适当 纤连蛋白的数量或类型将揭示有关其的信息 特定功能。 涉及诱变的更详细的研究 FN的特定部分将从两个FN的初步研究中遵循 同工型。