Genome-wide Dissection of the Retinal Determination Network

描述（由申请人提供）：本研究项目的目标是在全基因组范围内识别控制视网膜细胞发育的基因和遗传网络，这将为进一步研究奠定基础，旨在提高我们理解、诊断、治疗和预防人类视网膜疾病的能力。利用果蝇黑腹果蝇作为动物模型系统，利用遗传学、基因组学和计算生物学的组合方法，确定了由eye - eless （ey）调控的基因和途径，这是一种Pax6同源物，在控制视网膜细胞命运规范的遗传层次接近顶端。此外，这种新的组合方法一旦建立，可以应用于视网膜发育过程中其他转录因子的研究。作为Pax6的同系物，ey是一个主控基因，对眼睛发育既必要又充分。然而，目前只发现了ey的一个直接靶点，即眼正弦（so）。作为全面了解ey功能的重要一步，将使用三种独立的方法对其直接下游靶点进行全基因组筛选。基因表达谱以及计算方法已被用于鉴定数百个由ey调节的新候选基因。为了补充前两种方法，将进行染色质分析实验，以确定果蝇基因组中的Ey结合位点。进一步研究Ey的直接下游效应因子，以及本研究中发现的其他在不同遗传层次上起作用的新基因，将为我们全面了解视网膜发育的机制提供帮助。我们的具体目标是：1；果蝇视网膜发育过程中eye - eless直接下游靶点的鉴定果蝇基因组Ey结合位点的全基因组鉴定果蝇眼发育过程中新基因的功能表征本研究中发现的基因的研究将为更全面地了解果蝇的无眼功能和眼发育机制提供基础。由于ey及其许多下游基因在人类中高度保守，并且果蝇和哺乳动物视网膜之间存在许多发育相似之处，因此这项工作将直接影响我们对人类视网膜发育的理解。

DESCRIPTION (provided by applicant): The goal of this research project is to identify, genome-wide, the genes and genetic networks controlling retinal cell development, which will form the basis for further studies designed to improve our ability to understand, diagnose, treat and prevent human retinal diseases. Using the fruit fly Drosophila melanogaster as an animal model system, genes and pathways regulated by eyeless (ey), a Pax6 homolog that functions near the top of the genetic hierarchy controlling retinal cell fate specification will be identified using a combinatorial approach of genetics, genomics, and computational biology. In addition, this novel combinatorial approach, once established, can be applied to studies of other transcription factors during retinal development. A Pax6 homolog, ey functions as a master control gene that is both essential and sufficient for eye development. However, only one direct target of ey, sine oculis (so), has been identified. As an essential step toward a full understanding of ey function, a genome-wide screen for its direct downstream targets using three independent methods will be conducted. Gene expression profiles as well as computational approaches have been used to identify several hundred novel gene candidates regulated by ey. To complement the first two approaches, chromatin profiling experiments will be conducted to identify Ey binding sites in the Drosophila genome. Further studies of direct downstream effectors of Ey, as well as other novel genes acting at different levels of the genetic hierarchy identified in this study, will provide us with a comprehensive understanding of the mechanisms of retinal development. Our Specific Aims are: 1. Identification of direct downstream targets of Eyeless during Drosophila retinal development 2. Genome-wide identification of Ey binding sites in the Drosophila genome 3. Functional characterization of novel genes during Drosophila eye development Studies of genes identified in this study will provide the basis for a more complete understanding of eyeless function as well as mechanisms of eye development. Since both ey and many of its downstream genes are highly conserved in humans and many developmental parallels exist between Drosophila and mammalian retina, this work will directly impact our understanding of human retinal development.