FUNCTIONAL STUDY OF BRN-3B IN RETINAL GANGLION CELLS

描述（改编自申请人摘要）：该提案涉及 通过设计用于分析体外和体内的研究， Brn-3b，一种POU结构域转录因子，在RGC发育中的作用， 探索病理条件下RGC变性的潜在机制 使用brn-3b基因座作为分子工具的条件。 第一、 Brnb-3b的转录特性和结构/功能关系 将使用包括凝胶迁移率变化的生物化学方法进行分析， 甲基化干扰足印和共转染转录 测定。 这些体外研究将提供关于如何 Brn-3b可能特异性调控RGC中靶基因的表达 发展 其次，各种组织化学和免疫化学方法 Brn-3b是否在分裂中被激活 视网膜母细胞，并比较Brn-3b之间的细胞增殖和死亡 （+/+）和（-/-）视网膜。 这些体内研究预计将获得关键的 在RGC期间，关于Brn-3b功能的定时和模式的信息 发展 第三，Brb-3b将被同源重组取代 在小鼠中用碱性磷酸酶报告基因检查 轴突，或与bcl-2基因，以研究拯救细胞凋亡的Brn-3b表达 RGC。 所得数据将有助于理解Brn-3b的功能 在RGC中，也可能揭示病理性视网膜病变中RGC的变性。 条件 总的来说，对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，这些信息应该有助于更好地 了解导致某些致盲性疾病的机制，以及 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.