Brn-3 Transcription Factors in Retinal Development

视网膜发育中的 Brn-3 转录因子

• 批准号: 7060803
• 负责人: Lin Gan
• 金额: $ 38.08万
• 依托单位: UNIVERSITY OF ROCHESTER
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-05-01 至 2008-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7060803
• 关键词: DNA binding protein; apoptosis; cell differentiation; cell growth regulation; developmental genetics; gene deletion mutation; gene expression; genetically modified animals; laboratory mouse; neural degeneration; neurogenesis; neuronal guidance; neuroregulation; phenotype; polymerase chain reaction; protein sequence; retina; retinal ganglion; transcription factor; transfection

DESCRIPTION (provided by applicant): Retinal degeneration diseases pose great health risks. Currently, the effective cures and prevention methods remain elusive. Identification of genes and genetic processes important for the formation and survival of retinal neurons will lead to the development of prevention drugs targeting these genes and processes and to develop approaches to replace the degenerated retinal neurons by transplantation or regeneration from neuronal stem cells. The long-term objective of this proposal is to understand the role of Brn-3 POU-domain transcription factors in retinal neurogenesis and to identify and characterize the roles of their downstream effect genes. The three brn-3 genes, brn-3a, brn-3b and brn-3c, share a highly conserved functional POU-domain and their expression during neurogenesis and in adult is largely overlapping. Targeted mutagenesis studies in mice have shown that deletion of each brn-3 genes lead to unique neuronal phenotypes with the loss of a selected group of neurons. Intriguingly, the uniqueness of knockout phenotypes in each brn-3 mutant closely correlated to its distinctive spatiotemporal expression pattern. In retina, expression of brn-3 genes is mostly overlapping in retinal ganglion cells (RGCs) and the onset of brn-3b expression precedes those of brn-3a and brn-3c. Deletion of brn-3b results in the terminal differentiation failure and apoptosis of approximate 70% of RGCs. Loss of brn-3c has similar effects on a small percentage of RGCs. To further understand the roles of brn-3 genes in retinal neurogenesis and to explore the common molecular mechanisms of brn-3 function, in this application, we will: 1) use the transgenic approach to test the functional equivalence of brn 3 genes. The coding regions of brn-3a and brn-3c will be used to replace brn-3b in the knock-in experiments. The ability of knock-in brn-3 genes to rescue the retinal phenotypes associated with brn-3b knockout will be examined; 2) determine the role of brn-3a in the development of RGCs. Defects in retina of brn-3a-null mice or mice null for brn-3b and brn-3a will be analyzed; 3) identify and characterize in vivo the role of brn-3b downstream genes. We have shown that loss of Brn-3b leads to the diminished RGC expression of transcription factors Gfi-1 and LMO2. Gfi-1 is a zinc-finger protein required for the survival of inner ear hair cells. Recently, we have identified LMO2 as the first transcription factor expressed in the GCL of developing retina in a low-nasal-to-high-temporal gradient. Transgenic approaches will be used to investigate their roles in retinal development, particularly the axon pathfinding and survival of RGCs.

描述（由申请人提供）：视网膜变性疾病带来巨大的健康风险。目前，有效的治疗和预防方法仍然难以捉摸。对视网膜神经元形成和存活重要的基因和遗传过程的鉴定将导致针对这些基因和过程的预防药物的开发，并开发通过神经元干细胞移植或再生来替代退化视网膜神经元的方法。该提案的长期目标是了解 Brn-3 POU 结构域转录因子在视网膜神经发生中的作用，并识别和表征其下游效应基因的作用。三个 brn-3 基因 brn-3a、brn-3b 和 brn-3c 共享一个高度保守的功能性 POU 结构域，并且它们在神经发生期间和成人中的表达在很大程度上是重叠的。小鼠定向诱变研究表明，删除每个 brn-3 基因会导致独特的神经元表型，并导致选定的神经元组丢失。有趣的是，每个 brn-3 突变体的敲除表型的独特性与其独特的时空表达模式密切相关。在视网膜中，brn-3 基因的表达在视网膜神经节细胞 (RGC) 中大部分重叠，并且 brn-3b 表达的开始先于 brn-3a 和 brn-3c 的表达。 brn-3b 的缺失会导致约 70% 的 RGC 终末分化失败和凋亡。 brn-3c 的缺失对一小部分 RGC 也有类似的影响。为了进一步了解brn-3基因在视网膜神经发生中的作用，并探讨brn-3功能的常见分子机制，在本申请中，我们将：1）使用转基因方法测试brn 3基因的功能等价性。 brn-3a和brn-3c的编码区将在敲入实验中替代brn-3b。将检查敲入 brn-3 基因拯救与 brn-3b 敲除相关的视网膜表型的能力； 2)确定brn-3a在RGC发育中的作用。将分析 brn-3a 缺失小鼠或 brn-3b 和 brn-3a 缺失小鼠的视网膜缺陷； 3) 鉴定并表征brn-3b下游基因的体内作用。我们已经证明，Brn-3b 的缺失会导致转录因子 Gfi-1 和 LMO2 的 RGC 表达减少。 Gfi-1 是内耳毛细胞存活所需的锌指蛋白。最近，我们发现 LMO2 是第一个在发育中的视网膜 GCL 中以低鼻到高颞梯度表达的转录因子。转基因方法将用于研究它们在视网膜发育中的作用，特别是轴突寻路和 RGC 的存活。