GENOME WIDE ANALYSIS OF EPITHELIAL TUBE FUSION

肾小管网络的形成对肺、肾和血管系统等器官的形成至关重要。果蝇气管系统提供了一个很好的模型系统，其中管状网络的形成可以使用强大的遗传、分子和基因组工具进行研究。先前的研究表明，果蝇和脊椎动物在介导管分支的分子途径上有高度的保守性。这一建议侧重于气管网络形成的另一个方面，分支融合。生长中的分支必须识别并向适当的融合伙伴移动，然后对齐并形成适当的连接。这些融合事件对于形成相互连接的网络至关重要，但在细胞和分子水平上仍然知之甚少。融合事件是由一个被称为融合细胞的单个细胞介导的，它位于生长分支的末端。为了确定在分支融合过程中起作用的基因，融合细胞将通过全动物细胞分选（WACS）从胚胎中分离出来，由这些细胞制成的cdna将用于探测DNA微阵列。将融合细胞中表达的基因与其他气管细胞中表达的基因进行比较，可以鉴定融合细胞特异性基因。这些候选基因的表达模式将通过原位杂交验证，这些基因的功能表型损失将通过RNA干扰（RNAi）实验确定。基因将被分类并按路径排序，为融合过程中每个细胞步骤的详细分子理解奠定基础。

The creation of tubular networks is essential to the production of such organs as the lungs, the kidneys, and the vascular system. The Drosophila tracheal system provides an excellent model system in which the formation of tubular networks can be studied using powerful genetic, molecular, and genomic tools. Previous work has shown that there is a high degree of conservation between flies and vertebrates in the molecular pathways used to mediated tube branching. This proposal focuses on another aspect of tracheal network formation, branch fusion. Growing branches must identify and move towards the appropriate fusion partner, then align and form an appropriate connection. These fusion events are essential to the formation of an interconnected network, and remain poorly understood at the cellular and molecular level. Fusion events are mediated by a single cell, called a fusion cell, found at the ends of growing branches. To identify the genes that act during branch fusion, fusion cells will be isolated from embryos by whole animal cell sorting (WACS), and cDNAs made from these cells will be used to probe DNA microarrays. Comparison of genes expressed in fusion cells to genes expressed in other tracheal cells allow identification of fusion cell- specific genes. The expression pattern of these candidate genes will be verified by in situ hybridization, and the loss of function phenotypes of these genes will be determined by RNA interference (RNAi) experiments. Genes will be categorized and ordered into pathways, laying the foundation for a detailed molecular understanding of each cellular step in the fusion process.