FUNCTIONAL STUDY OF BRN-3B IN RETINAL GANGLION CELLS

描述(改编自申请人的摘要)：有关建议 通过设计在体外和体内分析的研究， BRN-3b，一个POU结构域转录因子，参与RGC的发育，并 病理状态下RGC退变的潜在机制探讨 以BRN-3b基因座为分子工具的条件。第一, Brnb-3b的转录特性及其结构/功能关系 将使用包括凝胶迁移率变化在内的生化方法进行分析， 甲基化干扰足印和共转染转录 化验。这些体外研究将提供有关如何 BRN-3b可能特异性调控其靶基因在RGC中的表达 发展。第二，各种组织化学和免疫化学方法 将用于调查BRN-3b是否在分割中被激活 视网膜母细胞，比较BRN-3b的细胞增殖和死亡 (+/+)和(-/-)视网膜。这些体内研究有望获得至关重要的 关于RGC期间BRN-3b功能的时间和模式的信息 发展。第三，BRB-3b将被同源重组所取代 在带有碱性磷酸酶报告器的小鼠中检查投射 轴突，或与bcl2基因联合研究BRN-3b表达中细胞凋亡的挽救 RGC。所得数据将有助于理解BRN-3b的功能 也可能为病理性视网膜节细胞的退行性变提供线索。 条件。总之，关于BRN-3b功能的拟议研究将提供 对控制哺乳动物的分子机制的基本见解 视网膜发育。从长远来看，这些信息应该会有更好的帮助 了解导致某些失明障碍的机制，并 可能为治疗方式提供新的方向。

DESCRIPTION (Adapted from applicant's abstract): The proposal is concerned with studies designed to analyze both in vitro and in vivo, the function of Brn-3b, a POU-domain transcription factor, in RGC development, and to explore underlying mechanisms for RGC degeneration under pathological conditions using the brn-3b gene locus as a molecular tool. First, transcriptional properties and structure/function relationships of Brnb-3b will be analyzed using biochemical approaches including gel mobility shift, methylation interference foot printing, and co-transfection transcription assays. These in vitro studies will provide initial information on how Brn-3b may specifically control its target gene expression in RGC development. Secondly, various histochemical and immunochemical approaches will be utilized to investigated whether Brn-3b is activated in dividing retinoblasts, and to compare cell proliferation and death between Brn-3b (+/+) and (-/-) retinas. These in vivo studies are expected to gain crucial information regarding the timing and mode of Brn-3b function during RGC development. Thirdly, Brb-3b will be replaced by homologous recombination in mice with the alkaline phosphatase reporter to examine the projections of axons, or with bcl-2 gene to study rescue of apoptosis in Brn-3b-expressing RGCs. The resulting data will help in understanding the function of Brn-3b in RGCs, and may also shed light on degeneration of RGCs in pathological conditions. Together, the proposed studies on Brn-3b function will provide fundamental insights into the molecular mechanism that control mammalian retinal development. In the long term, the information should help better understand the mechanisms leading to certain blinding disorders, and also may provide new directions for treatment modalities.