The Role of the Rx Homeobox Genes in Retinal Formation

描述（由申请人提供）：脊椎动物眼睛的发育是一个复杂的过程，需要几个关键转录因子的相互作用。其中之一是含有配对同源框的基因Rx。Rx属于一个小的保守基因家族，在所有脊椎动物物种中显示几乎相同的表达模式。我们已经发现Rx对于正常的眼睛发育是必不可少的，并且其错误表达对眼睛形态有深远的影响。注射Rx RNA的非洲爪蟾胚胎发育出异位视网膜组织，并在神经视网膜中显示过度增殖。携带该基因无效等位基因的小鼠胚胎不会形成视杯，因此不会发育眼睛。在这些胚胎中没有表达眼睛发育的标志物，这表明视网膜祖细胞在没有Rx功能的情况下不会形成。由于Rx是脊椎动物眼睛形成的关键组成部分，我们提出了与其在眼睛发育中的功能相关的四个具体目标： 具体目标1：分析缺乏Rx功能的细胞对正常视网膜结构的贡献能力。为此，我们将从Rx-/-和野生型细胞中制备嵌合小鼠胚胎。该实验将确定Rx-/-细胞是否可以在野生型细胞存在下促进正常的眼睛发育。 具体目标2：确定调控Rx基因时空表达的分子机制。Rx调控元件将使用报告构建体鉴定。 具体目标3：FGF信号传导对视网膜细胞的特化和存活的影响。为此目的，显性负性FGF受体将使用Rx调节序列在发育中的视网膜中表达。将监测FGF信号传导的消除对视网膜细胞类型发育的影响。 具体目标4：操纵非洲爪蟾视网膜中的基因表达，以研究发育和分化。这将通过使用Rx调节区在转基因非洲爪蟾的视网膜祖细胞中靶向表达特定蛋白质来实现。将特别关注Otx 2和Xbarl含同源框的基因。

DESCRIPTION (provided by applicant): Development of the vertebrate eye is a complex process that requires interaction of several key transcription factors. One of them is the paired-like homeobox-containing gene Rx. Rx belongs to a small, conserved family of genes that displays an almost identical expression pattern in all of the vertebrate species. We have found that Rx is essential for normal eye development and its misexpression has profound effects on eye morphology. Xenopus embryos injected with Rx RNA develop ectopic retinal tissue and display hyperproliferation in the neuroretina. Mouse embryos carrying a null allele of this gene do not form optic cups and consequently do not develop eyes. No markers of eye development are expressed in these embryos suggesting that retinal progenitor cells do not form in the absence of Rx function. Since Rx is a key component in formation of vertebrate eyes, we propose four specific goals related to its function in eye development: Specific Aim 1: Analysis of the ability of cells lacking Rx function to contribute to normal retinal structures. For this purpose, we will make chimeric mouse embryos from Rx-/- and wild type cells. This experiment will determine whether Rx-/- cells can contribute to normal eye development in the presence of wild type cells. Specific Aim 2: Identification of the molecular mechanism that regulates the spatial and temporal expression of Rx genes. Rx regulatory elements will be identified using reporter constructs. Specific Aim 3: Effect of FGF signaling on the specification and survival of retinal cells. For this purpose dominant negative FGF receptors will be expressed in the developing retina using Rx regulatory sequences. Effects of elimination of FGF signaling on the development of retinal cell types will be monitored. Specific Aim 4: Manipulation of gene expression in the Xenopus retina in order to study development and differentiation. This will be achieved by a targeted expression of specific proteins in the retinal progenitor cells of transgenic Xenopus using the Rx regulatory region. Specific attention will be paid to the Otx2 and Xbarl homeobox-containing genes.