BRN-3 POU DOMAIN PROTEINS IN RETINAL DEVELOPMENT

描述（由申请人提供）：本项目的长期目标 应用程序是获得深入了解分子事件，导致 哺乳动物视网膜神经节细胞的分化。视网膜神经节细胞 对正常视力至关重要，它们的丧失会导致严重的眼部疾病， 人类例如，眼睛内的眼内压升高可以触发 神经节细胞凋亡增强，进而导致青光眼。尽管 然而，他们的重要性，视网膜相关基因的知识基础， 神经节细胞的形成和存活是基本的。在本申请中， 实验提出，重点对Brn-3b，POU结构域转录 因子，在视网膜神经节细胞中。在小鼠中，brn-3b基因是 有丝分裂后祖细胞作为神经节细胞激活的第一基因 分化开始了。尽管这种早期激活，brn-3b不是 神经节细胞的初始规格所需的，但它是必不可少的 正常的分化和存活。具有靶向缺失的小鼠， BRN-3B具有缺陷性视网膜，损失了大部分神经节细胞。因此，brn-3b 是一个关键基因，标志着对神经节细胞命运的承诺， 对视网膜神经节细胞的存活至关重要。前两个目标 本申请使用BRN-3B基因座来探测视网膜神经节的早期事件 细胞形成最后两个目标涉及的转录特性， Brn-3b具体目的：（1）检验视网膜病变的假设， 当神经节细胞形成时，它们抑制它们的进一步产生。拟议 实验将专门消融视网膜神经节细胞， 靶向白喉毒素;（2）确定内的顺式调节元件 在有丝分裂后激活brn-3b brn-3b转录控制区 神经节细胞祖细胞brn-3b内的顺式调节元件 将使用BAC转基因技术鉴定转录控制区。 （3）研究Brn-3b在视网膜神经元中的功能特异性 神经节细胞分化实验将采用HSV介导的基因 转移到培养的视网膜外植体中;（4）确定brn-3b是否可以 在将祖细胞转化为视网膜神经节细胞命运中起作用。Math5 是视网膜神经节细胞形成所需的前神经bHLH基因。BRN-3b 将在math 5位点被错误表达，以确定brn-3b是否 足以促进神经节细胞分化， 数学5

DESCRIPTION (provided by applicant): The long-term objective of this application is to gain insights into the molecular events that lead to the differentiation of mammalian retinal ganglion cells. Retinal ganglion cells are essential for normal vision and their loss contributes to major eye diseases in humans. For example, elevated intraocular pressure within the eye can trigger enhanced apoptosis in ganglion cells, which in turn leads to glaucoma. Despite their importance, however, the knowledge base of genes associated with retinal ganglion cell formation and survival is rudimentary. In this application, experiments are proposed that focus on Brn-3b, a POU-domain transcription factor, in retinal ganglion cells. In the mouse, the brn-3b gene is among the first genes activated in postmitotic progenitor cells as ganglion cell differentiation begins. In spite of this early activation, brn-3b is not required for the initial specification of ganglion cells, but it is essential for their normal differentiation and survival. Mice with targeted deletions in brn-3b have defective retinas with a loss of most ganglion cells. Thus, brn-3b is a critical gene that marks the commitment to a ganglion cell fate and is essential for the survival of retinal ganglion cells. The first two aims of this application use the brn-3b locus to probe early events of retinal ganglion cell formation. The final two aims concern the transcriptional properties of Brn-3b. The Specific Aims will: (1) Test the hypothesis that as retinal ganglion cells form, they inhibit their further production. The proposed experiments will specifically ablate retinal ganglion cells using genetically targeted diphtheria toxin; (2) Identify the cis-regulatory elements within the brn-3b transcriptional control region that activate brn-3b in postmitotic ganglion cell progenitors. The cis-regulatory elements within the brn-3b transcriptional control region will be identified using BAC transgenic analysis; (3) Investigate the functional specificity of Brn-3b in retinal ganglion cell differentiation. The experiments will employ HSV-mediated gene transfer into cultured retinal explants; (4) Determine whether brn-3b can function in committing progenitor cells to a retinal ganglion cell fate. Math5 is a proneural bHLH gene required for retinal ganglion cell formation. Brn-3b will be misexpressed at the math5 locus to determine whether brn-3b is sufficient to promote ganglion cell differentiation in the presence and absence of math5.