Investigation of Mammalian Retinal Neuron Development

描述（由申请人提供）：本提案使用小鼠模型研究人类眼部疾病的根本原因。我们专注于Notch信号通路，这是在多种哺乳动物组织中至关重要的。具体而言，Notch信号传导调节增殖、凋亡、细胞形状变化、分化和干细胞维持。该提案中的实验将1）阐明视网膜神经节细胞（RGC）神经发生期间Notch信号传导与Math 5之间的上位关系，2）探索Rbpj的多种视网膜神经元表型，以及3）定义视网膜神经发生期间Notch配体Deltalike 1的需求。由于Notch信号在发育过程中被广泛使用，因此大多数Notch途径基因中的小鼠突变已经产生。使用靶向缺失小鼠（完全突变和条件等位基因），我们建议了解视网膜神经节细胞和视锥和视杆感光细胞形成过程中对经典Notch信号的要求。一些研究将采用条件（cre-lox）小鼠品系、组织学、免疫组织化学、原位杂交、小鼠胚胎学和PCR基因分型。其他人将使用人类视网膜母细胞系在体外测试调控关系。这些研究将为所有后生动物发育的基础--生长、形态发生和分化过程提供新的信息。这项工作将产生一个更好的理解锥杆营养不良，视神经发育不全，发育不全，以及有助于视网膜细胞发育的基本机制与直接相关的基因或细胞为基础的视网膜治疗。这些发现对癌症生物学领域也有广泛的应用价值，因为Notch 1或Hes 1的过度活化表达存在于多种人类肿瘤中。公共卫生相关性：本研究的目的是了解哺乳动物视网膜神经元形成的潜在分子机制，使用小鼠模型。我们建议通过研究视网膜形成的哪些方面需要Notch细胞间信号通路，以及它如何调节Math 5 bHLH转录因子来实现这一点。在身体的其他部位，Notch信号控制细胞的形状变化，生长和死亡。由于这些原因，Notch途径的突变可能导致癌症。对Notch如何、何时、何地在视网膜中起作用的透彻理解只是表面上的。这些研究将在单细胞水平上对视网膜如何发育提供更深入的了解，并有助于更好地设计疾病治疗方法，如Leber's congeital amaurosis，锥-杆营养不良，色盲，视神经发育不全/发育不全和青光眼。

DESCRIPTION (provided by applicant): This proposal investigates the underlying causes of human ocular diseases using mouse models. We focus on the Notch signaling pathway, which is critically required in multiple mammalian tissues. In particular, Notch signaling regulates proliferation, apoptosis, cell shape changes, differentiation and stem cell maintenance. Experiments in this proposal will 1) elucidate the epistatic relationship between Notch signaling and Math5 during retinal ganglion cell (RGC) neurogenesis 2) explore the multiple retinal neuron phenotypes of Rbpj, and 3) define the requirements of the Notch ligand Deltalike1 during retinal neurogenesis. Because Notch signaling is widely employed during development, mouse mutations in most Notch pathway genes have already been created. Using targeted deletion mice (wholly mutant and conditional alleles), we propose to understand the requirements for canonical Notch signaling during retinal ganglion cell and cone and rod photoreceptor formation. Some studies will employ conditional (cre-lox) mouse strains, histology, immunohistochemistry, in situ hybridization, mouse embryology and PCR genotyping. Others will test regulatory relationships in vitro using a human retinoblast cell line. These studies will contribute new information to the processes of growth, morphogenesis and differentiation, which are fundamental to all metazoan development. This work will yield a better understanding of cone-rod dystrophies, optic nerve aplasia, hypoplasia, as well as contribute to basic mechanisms of retinal cell development with direct relevany to gene- or cell-based retinal therapies. Findings here will also be widely useful to the field of cancer biology, since excess activated Notch1 or Hes1 expression occurs in a variety of human tumors. PUBLIC HEALTH RELEVANCE: The goal of this study is to understand the underlying molecular mechanisms of how mammalian retinal neurons forms, using mouse models. We propose to do this by investigating which aspects of retinal formation require the Notch cell-to-cell signaling pathway, and how it regulates the Math5 bHLH transcription factor. In other parts of the body, Notch signaling controls cell shape changes, growth, and death. For these reasons, mutations in the Notch pathway can cause cancer. A thorough understanding of how, when and where Notch acts in the retina has only been addressed superficially. These studies will provide deeper understanding, at the single cell level, of how the retina develops and contribute to the better design of disease therapies for diseases such as Leber's congeital amaurosis, cone-rod dystrophy, color blindness, optic nerve aplasia/hyplasia and glaucoma.