CDX-1 AND CDX-2 IN DEVELOPMENT

指导肠道发育的分子机制还没有被发现。 充分阐明。 同源结构域转录因子已被证明 对许多生物体的发育至关重要。初步 数据显示，mCdx-2可以激活肠道特异性基因，因此， 两个肠特异性小鼠尾相关同源结构域研究 家族基因mCdx-1和mCdx-2可能提供有关以下方面的重要信息： 控制肠道的分化和发育。客观 这一建议的特点是时间和细胞 这些Cdx基因的表达模式，并确定它们在 发育中的老鼠为实现这一目标，将实现以下具体目标： 1）在大肠杆菌中mCdx-1和mCdx-2表达的表征 通过原位杂交和免疫组织化学， 强调其细胞定位沿着纵向和 肠道的垂直轴（隐窝-绒毛轴）。2)的 评价Cdx基因在发育中的功能作用， 功能获得和功能丧失研究。 检查…的功能 mCdx-1和mCdx-2基因，功能获得性实验将 使用转基因小鼠进行。这些小鼠将被设计，使用 绒毛蛋白启动子，具有Cdx基因的异位表达， 内胚层来源的组织以及早熟表达。小鼠 将分析可能发生在肠内胚层的变化 在发育过程中，直接 组织学检查、原位杂交和免疫组织化学。 还将使用转基因小鼠进行功能丧失实验 与mCdx-2的显性负性蛋白突变体一起繁殖， 阻断天然蛋白质的结合。支配结构 负突变体将在绒毛蛋白启动子的控制下， 表达至内胚层来源的组织。转基因小鼠将被 分析了组织学和生理学变化， mCdx-2功能。 这些实验旨在阐明一些机制， 直接肠道发育和分化。的完成 本建议书的第一个目标是提供描述性信息， 将被用来分析潜在的变化，发生增加或 降低转基因小鼠中Cdx基因的功能。使用 转基因动物允许在体内研究去调节 基因，在这种情况下是转录因子mCdx-1和mCdx-2。的 理解转录调控的基本机制， 肠基因，通过研究尾部相关的同源结构域基因， 作为进一步了解肠道发育的一种手段， 可能会让我们深入了解影响消化系统疾病的过程。

The molecular mechanisms that direct intestinal development have not been fully elucidated. The homeodomain transcription factors have been shown to be of central importance for development in many organisms. Preliminary data shows that mCdx-2 can activate intestinal specific genes, therefore the study of two intestine specific mouse caudal-related homeodomain family genes, mCdx-1 and mCdx-2, may give essential information regarding the control of intestinal differentiation and development. The objective of this proposal is to characterize both the temporal and cellular patterns of expression of these Cdx genes and to determine their role in developing mice. To reach this objective the following specific aims will be pursued: l) The characterization of mCdx-l and mCdx-2 expression in developing mice by in situ hybridization and immunohistochemistry with an emphasis on their cellular localization along the longitudinal and vertical (crypt-villus axis) axes of the intestinal tract. 2) The evaluation of the functional role of Cdx genes in development by both gain-of-function and loss-of-function studies. To examine the function of the mCdx-l and mCdx-2 genes, gain-of-function experiments will be performed using transgenic mice. These mice will be designed, using the villin promotor, to have both ectopic expression of the Cdx genes in endodermally derived tissue as well as precocious expression. The mice will be analyzed for changes that may occur at the endoderm-intestinal transition and at the crypt-villus axis during development by direct histologic inspection, in situ hybridization and immunohistochemistry. Loss-of-function experiments will also be performed using transgenic mice bred with a dominant negative protein mutant of mCdx-2 that effectively blocks the binding of the native protein. The construct of the dominant negative mutant will be under the control of the villin promotor to direct expression to endodermally derived tissue. The transgenic mice will be analyzed for histologic and physiologic changes that occur with the loss of mCdx-2 function. These experiments are designed to elucidate some of the mechanisms that direct intestinal development and differentiation. The completion of the first objective in this proposal will give descriptive information which will be used to analyze potential changes that occur with increased or decreased function of the Cdx genes in the transgenic mice. The use of transgenic animals allows for an in vivo study into the deregulation of genes, in this case the transcription factors mCdx-l and mCdx-2. The understanding of the basic mechanisms of transcriptional regulation of intestinal genes, by studying the caudal related homeodomain genes, will serve as a means to further our knowledge of intestinal development and may give insight into the processes influencing digestive diseases.