MOLECULAR GENETICS OF A LENS INTRINSIC MEMBRANE PROTEIN

描述：（研究者摘要）。眼透镜是无血管的半固体器官，其功能是将入射光反射到视网膜上。伴随着该功能，透镜必须保持透明。透镜透明度的丧失通常是永久性的，称为白内障。细胞和器官功能，例如获得维持健康透明的透镜状态所必需的营养物，用于处理细胞废物去除，以及用于维持整体膜稳定性，通过选择和特定的蛋白质或透镜细胞膜内的蛋白质组来进行。适当控制这些蛋白质的表达及其随后的功能对于透镜的存活和透明度的维持是必不可少的。目前尚不清楚调节透镜中这些膜蛋白的机制。本研究的目的是了解透镜纤维细胞膜蛋白MP19基因（LIM2）的调控。为了实现这一目标，P.I.将研究以下主题：a）。研究小鼠To白内障突变体和人类先天性核性白内障及其与透镜膜LIM2基因的关系。利用转基因小鼠技术模拟To白内障，研究MP19的功能。B）。在瞬时转染试验中使用CAT报告基因构建体确定负责人LIM2基因（顺式元件）转录活性的区域。（c）。鉴定蛋白质：DNA相互作用（反式因子）与LIM2基础启动子区和上游顺式元件使用凝胶迁移率变化和DNA足迹程序。d）。研究LIM2上游区域的甲基化模式，以确定关键顺式元件的甲基化是否控制LIM2基因的功能。这些研究将使我们更好地了解MP19在维持透镜透明度中的作用及其在白内障形成中的可能作用。

DESCRIPTION: (Investigator's Abstract). The ocular lens is an avascular, semisolid organ whose function is to refract incoming light upon the retina. Concomitant with this function is a necessity for the lens to remain transparent. A loss of transparency of the lens is usually permanent and is termed a cataract. Cell and organ functions, such as obtaining nutrients necessary for maintaining the healthy, transparent lens state, for handling of cell waste removal, and for maintenance of overall membrane stability are carried out through select and specific proteins or groupings of proteins within the lens cell membrane. Proper controls on expression of these proteins and their subsequent function is essential for lens survival, and maintenance of transparency. The mechanisms regulating these membrane proteins in the lens are presently unknown. The goal of this project is to understand the regulation of the lens fiber cell membrane protein MP19 gene (LIM2). To achieve this goal, the P.I. will investigate the following topics: a). To characterize the mouse To cataract mutant and the human congenital nuclear cataract and their relationship to the lens membrane LIM2 gene. Using transgenic mouse technology to mimic the To cataract in order to study the function of MP19. b). To determine the region(s) responsible for transcriptional activity of the human LIM2 gene (cis elements), using CAT reporter gene constructs in transient transfection assays. c). Identify protein:DNA interactions (trans factors) associated with the LIM2 basal promoter region and upstream cis elements using gel mobility shift and DNA foot printing procedures. d). To investigate the methylation patterns of LIM2 upstream regions to determine if methylation of critical cis elements serves to control LIM2 gene function. These studies will give a better understanding of the role of MP19 in the maintenance of lens transparency and its possible role in cataract formation.