Genome-wide Dissection of the Retinal Determination Network

描述(申请人提供)：这项研究项目的目标是在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，这将成为进一步研究的基础，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，鉴定由眼盲(EY)调控的基因和途径，EY是控制视网膜细胞命运指定的遗传层次结构中接近顶端的Pax6同源基因。此外，这种新的组合方法一旦建立，就可以应用于视网膜发育过程中其他转录因子的研究。一个Pax6同源基因，ey起着主控基因的作用，这对眼睛的发育是必要的，也是足够的。然而，EY的直接靶点只有一个，即正弦眼球(Sine Oculis，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在全基因组范围内筛选其直接下游靶标。基因表达谱和计算方法已经被用来识别数百个受EY调控的新基因候选。为了补充前两种方法，将进行染色质图谱实验，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，以及本研究确定的在不同遗传层次上作用的其他新基因，将使我们对视网膜发育的机制有一个全面的了解。我们的具体目标是：1.在果蝇视网膜发育过程中鉴定失眼基因的直接下游靶点2.全基因组范围内鉴定果蝇基因组中的Ey结合位点3.果蝇眼睛发育过程中新基因的功能特征研究本研究确定的基因将为更全面地了解果蝇眼睛发育的功能和机制提供基础。由于EY及其下游基因在人类中高度保守，而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.