A novel strategy for identifying genes that regulate eye development

描述(申请人提供)：视网膜等组织的发育和维持取决于对细胞增殖、细胞存活和细胞与其邻近细胞之间适当的黏附相互作用的适当调控。虽然我们对其中许多过程的遗传调控的理解已经取得了很大的进步，但对果蝇成像盘的研究突出了细胞与细胞相互作用模式的存在，我们仍然不知道这些模式的存在。例如，对细胞竞争现象的研究表明，细胞能够评估其邻居的适合性，从而影响其增殖或生存。我们还知道，细胞似乎优先附着于来自成像盘相同部分的其他细胞，而避免与来自相同原基其他部分的细胞相互作用。这种偏好可能是绝对的，例如腿和翼盘的前后细胞明显分离，也可能是相对的，例如在发育中的果蝇眼中，克隆边界倾向于沿赤道下降。虽然细胞竞争和细胞划分都是在近40年前被发现的，但我们对这两种现象的机制基础仍然没有清楚的了解。因此，关于细胞如何在发育中的视网膜等组织中与邻近细胞相互作用，可能还有很多有待发现的地方。果蝇眼睛中的镶嵌屏幕已经被用来发现许多调控细胞增殖、细胞命运指定和光感受器分化的基因。重要的是，其中一些基因的突变引起的表型只有在椎间盘由两种不同类型的细胞组成的情况下才明显：野生型和突变型。在这些条件下，细胞增殖的变化表现为两种细胞群的相对代表性的变化，而不一定影响眼睛的大小，细胞-细胞亲和力的变化可以导致两种细胞群之间形成平滑的边界，而不是不规则的边界。虽然基于RNA介导的干扰(RNAi)的筛选在识别许多生物过程中的基因方面已经非常有效，但目前我们很难在镶嵌筛选的背景下使用这种方法。我们已经开发了一种新的方法CoinFLP，它允许我们使用RNAi在果蝇的眼睛中进行马赛克筛选，并通过一个试点筛选验证了这种方法，该筛选已经确定了几个以前与眼睛发育无关的基因。我们建议使用这种方法来识别和表征在发育中的果蝇眼睛中调节细胞增殖和细胞亲和力的基因。

DESCRIPTION (provided by applicant): The development and maintenance of a tissue such as the retina is contingent upon the proper regulation of cell proliferation, cell survival and appropriate adhesive interactions between cells and their neighbors. While great advances have been made in our understanding of the genetic regulation of many of these processes, studies in Drosophila imaginal discs have highlighted the existence of modes of cell-cell interactions that we still do not understand. For instance, studies of the phenomenon of cell competition have demonstrated that cells are capable of assessing the fitness of their neighbors and subsequently influencing their proliferation or survival. We also know that cells appear to preferentially adhere to other cells from the same portion of the imaginal disc while avoiding interactions with cells from other parts of the same primordium. This preference may be absolute such as the clear separation of anterior and posterior cells in leg and wing discs or relative, such as the tendency of clonal boundaries to fall along the equator in the developing Drosophila eye. While both cell competition and compartmentalization were discovered almost forty years ago, we still do not have a clear understanding of the mechanistic basis of either phenomenon. It is therefore likely that much remains to be discovered about the ways in which cells interact with the neighbors in tissues such as the developing retina. Mosaic screens in the Drosophila eye have been used to discover many genes that function in regulating cell proliferation, cell fate specification and photoreceptor differentiation. Importantly, the phenotypes elicited by mutations in some of these genes are only obvious in a situation where the disc is composed to two different types of cells: wild type and mutant. Under these conditions, changes in cell proliferation manifest as a change in the relative representation of the two populations of cells without necessarily affecting eye size and changes in cell-cell affinity can result in the formation of smooth, rather than irregular boundaries between the two populations. While screens based on RNA-mediated interference (RNAi) have been extremely effective in identifying genes in a number of biological processes, it is currently difficult to us this approach in the context of a mosaic screen. We have developed a new method, CoinFLP, which allows us to conduct mosaic screens in the Drosophila eye using RNAi and have validated this approach with a pilot screen that has already identified several genes not previously implicated in eye development. We propose to use this approach to identify and characterize genes that regulate cell proliferation and cell affinity in the developing Drosophila eye.