EXPRESSION AND ROLE OF T-BOX GENES IN MOUSE DEVELOPMENT

拟议研究的长期目标是描述 一个新发现的古老的胚胎表达家族的成员 小鼠基因，T-box基因，并确定它们的功能作用 在胚胎发育中发挥作用。这些基因共享一个保守的质子基序， 与DNA结合活性共定位的T-box 小鼠的典型T基因座，一个调节中胚层的基因座 队形。因为广域之间的进化守恒 从线虫到老鼠的不同物种，这些基因可能会发挥作用 在所有脊椎动物的发育过程中起着重要作用。我们的初步结果是 6个新发现的T-box基因揭示了时间和空间表达 在胚胎发育过程中的模式，一些独特的和一些重叠的 建议这些基因在不同方面发挥交互作用 发展。我们将详细说明这些基因的表达，并关键地 使用突变分析测试它们的功能角色。 尽管这项研究有具体的目标，将为 已经确定的单个基因，总体目标要宽泛得多 在范围内，并包含了试图理解 基因家族的各种成员，以及进化的基础 共同主题的保护。这是我们的信念，基于我们的表达 数据，并通过与其他具有保守、共享基序的基因家族的类比 例如Hox和Pax，T-box基因家族编码一组DNA 结合蛋白在发育过程中扮演着不同的角色 发信号。我们的目的是在我们的研究中增加新的T-box基因 它们被识别出来，并集中精力研究单个基因或 在阐明过程中似乎信息最丰富的一组相关基因 发育机制。为了实现这个更大的目标，这个 建议书是由调查员发起的两部分互动的一个组成部分 研究项目补助金(IRPG)。我们的合作者，李·西尔弗博士 普林斯顿大学已经得到资助，在他的 实验室。本建议书的提交反映了我们共同的 目标、互补专业知识和意图继续我们的 现有的合作安排来调查这个古老的家族 老鼠的基因。 具体目的1.确定小鼠T-box基因的表达模式，1- 6、胚胎发生、器官发生和成体发育。我们将使用 组织切片和整体的Northern印迹分析、原位杂交 以及基因靶向构建体，该构建体包括由 内源性启动子。 具体目的2.通过创建空基因来确定T-box基因的功能 胚胎干细胞中使用同源重组的突变。这个 将对产生的突变动物的表型进行分析，突变动物将 交配在一起研究不同基因之间可能的相互作用。

The long-term objectives of the proposed research are to characterize the members of a newly discovered, ancient family of embryonically-expressed mouse genes, the T-box genes, and to determine the functional roles they play in embryonic development. These genes share a conserved proton motif, the T-box, which co-localizes with the DNA binding activity of the prototypical T locus of the mouse, a locus which regulates mesoderm formation. Because of the evolutionary conservation between widely divergent species from nematode to mouse, these genes are likely to play a role in the development of all vertebrates. Our preliminary results with 6 newly discovered T-box genes reveal temporal and spatial expression patterns during embryogenesis, some unique and some overlapping that suggest interactive roles for these genes in different aspects of development. We will detail the expression of these genes and critically test their functional roles using mutational analysis. Although the research has specific goals that will be accomplished for the individual genes already identified, the overall objective is much broader in scope, and encompasses an attempt to understand the relationship between the various members of the gene families, and the basis of the evolutionary conservation of the shared motif. It is our belief, based our expression data and by analogy with other gene families with conserved, shared motifs such as Hox and Pax, that the T-box family of genes encodes a set of DNA binding proteins that each play a distinctive role in developmental signaling. our intention is to add new T-box genes to our investigation as they are identified and to concentrate efforts on individual genes or groups of related genes that appear most informative in elucidating developmental mechanisms. In order to accomplish this larger aim, this proposal is one component of a two-part Investigator-Initiated Interactive Research Project Grant (IRPG). Our collaborator, Dr. Lee Silver of Princeton University has been funded to pursue complementary work in his laboratory. The submission of this proposal as an IRPG reflects our shared objectives, complementary expertise and intention of continuing our existing collaborative arrangement to investigate this ancient family of mouse genes. Specific Aim 1. Determine the expression pattern of mouse T-box genes, 1- 6, during embryogenesis, organogenesis and in the adult. We will use Northern blot analysis, in situ hybridization of tissue sections and whole mounts, and gene targeting constructs that include a marker gene driven by the endogenous promoter. Specific Aim 2. To determine the function of T-box genes by creating null mutations using homologous recombination in embryonic stem cells. The phenotype of resulting mutant animals will be analyzed and the mutants will be mated together to study possible interactions between different genes.