Pros/Prox1 and Lens Development

描述(申请人提供)：在脊椎动物中，功能性晶状体需要晶状体纤维细胞分化、延长和表达大量适当的晶状体蛋白分子。影响其中任何一个过程的因素都是有害的，并可能导致白内障疾病的形成。基因敲除研究表明，小鼠转录因子Proxl调节晶状体纤维的伸长和晶体蛋白基因的表达。不幸的是，Prox1基因敲除的动物在胚胎13.5天前死亡，这使得进一步分析这一表型变得困难。我们的初步数据表明，果蝇Proxl的直系同源基因，称为Propero(Pros)，也是晶状体形成所必需的。与Proxl类似，Pros影响晶状体细胞分化和晶状体蛋白基因表达。这是无脊椎动物和脊椎动物晶状体发育共享相似转录调控途径的首批报告之一。有趣的是，Pros和Prox1在晶状体细胞分化过程中也表现出核和细胞质的表达，这表明这些蛋白也具有非转录作用。这些研究旨在探索Pros/Prox1介导的晶状体发育的进化保守基础。为此，我们将：1)更准确地描述Pros在晶状体细胞发育中的作用，2)测试Pros在晶状体发育中的细胞内定位，以及3)用Pros/Prox1转录因子家族表征功能保守性。这些研究将结合体外生化和转录调节分析、定点突变和体内遗传分析，为Pros/Prox1蛋白在晶状体发育过程中的功能提供广阔的前景。这项工作将提供必要的框架，以进行更深入的基因研究，旨在确定晶状体正常发育所必需的关键调控途径，以此作为一种手段，了解患有先天性和年龄相关性白内障的患者这些途径的哪些方面受到干扰。Laye：白内障是由于我们的晶状体没有得到适当的发育或维持而导致的。不幸的是，人们对正常晶状体是如何发育的知之甚少，这使得我们很难设计出防止晶状体形成异常的方法，比如在白内障疾病中。这项建议的重点是开发一种新的遗传模型，使我们能够快速确定晶状体发育所必需的新途径，以此作为绕过导致白内障形成和失明的事件的一种手段。

DESCRIPTION (provided by applicant): In vertebrates, a functional lens requires the ability of lens fiber cells to differentiate, elongate, and express abundant levels of the appropriate crystallin molecules. Factors which affect any of these processes are detrimental and can lead to the formation of cataract disease. Knockout studies have shown that the mouse transcription factor Proxl regulates lens fiber elongation and crystallin gene expression. Unfortunately, Proxl knockout animals die by embryonic day 13.5, making further analysis of this phenotype difficult. Our preliminary data demonstrate that the Drosophila ortholog of Proxl, called Prospero (Pros), is also necessary for lens formation. Similar to Proxl, Pros affects lens cell differentiation and crystallin gene expression. This is one of the first reports that lens development in invertebrates and vertebrates shares similar transcriptional regulatory pathways. Interestingly, Pros and Proxl also exhibit both nuclear and cytoplasmic expression during lens cell differentiation, suggesting a non-transcriptional role for these proteins as well. The studies outlined here are aimed to explore the evolutionary conserved basis for Pros/Prox1-mediated lens development. For this, we will: 1) more precisely characterize the role for Pros in lens cell development, 2) test the contribution of Pros intracellular localization in lens development, and 3) characterize functional conservation with the Pros/Prox1 family of transcription factors. These studies will use a combination of in vitro biochemical and transcriptional regulatory assays, site-directed mutagenesis, and in vivo genetic analyses to provide a broad perspective on how Pros/Prox1 proteins function during lens development. This work will provide the framework necessary to perform more in-depth genetic studies aimed at identifying key regulatory pathways necessary for normal lens development as a means for understanding what aspects of these pathways are disturbed in patients suffering from congenital and age- related cataracts. LAY: Cataracts result from the failure of our lens to properly develop or to be maintained. Unfortunately, not much is known about how a normal lens develops, making it difficult for us to devise ways to prevent abnormal lens formation such as in cataract disease. This proposal is focused on developing a new genetic model that will allow us to rapidly identify new pathways that are necessary for lens development as a means to circumvent events that lead to cataract formation and blindness.