Genetic proteomic and functional analysis of junctional complexes in Drosophila

我们的身体依赖于屏障上皮的完整性；屏障上皮是将我们与外界隔开的细胞片(如皮肤、肠道、肺)，或将我们身体的不同部位分开。这类上皮中的数千个细胞具有高度专业化的连接，将它们拉紧在一起，以防止泄漏(紧密的上皮)或仅允许特定溶质的流动(泄漏的上皮)。这种完整性的问题可能是严重的或致命的(例如腹膜炎、多发性硬化症、肾脏疾病)。是什么让结点变紧或漏水？在连接处发现的许多蛋白质都是已知的。然而，除了一些人类遗传病和少数基因敲除的小鼠品系外，很难区分出每种蛋白质的作用。我们建议研究模式昆虫黑腹果蝇肾脏中的隔状连接(脊椎动物“紧密”连接的同源物)。这是一个理想的系统，因为我们可以廉价而快速地干预我们选择的细胞中的基因功能。我们将使用蛋白质组学来鉴定连接中的主要蛋白质，然后突变蛋白质组结果所涉及的基因，并研究连接是保持紧密还是变得漏水。这将使人们能够全面研究组成连接的蛋白质的相对作用。

Our bodies rely on the integrity of our barrier epithelia; the sheets of cells (like skin, gut, lung) that separate us from the outside world, or keep different regions of our bodies apart. The thousands of cells in such epithelia have highly specialised junctions that zip them together, in order to prevent leakage (a 'tight' epithelium) or to permit flux of only specific solutes (a 'leaky' epithelium). Problems with this integrity can be serious or fatal (e.g. peritonitis, multiple sclerosis, kidney disease). What makes a junction either tight or leaky? Many of the proteins that are found in junctions are already known. However, with the exceptions of some human genetic diseases and a few 'knockout' mouse lines, it is hard to tease apart the contributions of each protein. We propose to study the septate junction (the homologue of the vertebrate 'tight' junction) in the kidney of the model insect, Drosophila melanogaster. This is an ideal system, because we can cheaply and quickly intervene in gene function in cells of our choice. We will use proteomics to identify the major proteins in the junction, then mutate the genes implicated by the proteomics results, and study whether the junctions remain tight, or become leaky. This will allow a comprehensive study of the relative roles of the proteins that make up the junction.