Acquisition of a Drosophila chamber for culturing Drosophila strains

这项建议是收购一个果蝇室，以取代一个旧的和破碎的 室，这是用来培养果蝇菌株在我们的实验室。美国NIH R15 研究项目是研究一个知之甚少的Osiris（Osi）基因家族在调节 在果蝇气管管成熟，这是一个首要的系统，以揭示基本的 管状器官形成的机制。输卵管畸形导致各种人类疾病， 如多囊肾和血管疾病。 果蝇的气管是一个由上皮细胞构成的分支网络， 细胞围绕顶端腔。在管成熟过程中，顶端分泌物爆发沉积 大量的腔基质成分到顶端细胞外腔。然后，这个流明 在空气可以填充管腔之前被清除。先前的研究表明，蛋白质的成分 运输途径与管腔蛋白的分泌/清除有关。的 获奖项目的目标是揭示Osi蛋白作为“更广泛的协调者”的功能。 蛋白质运输过程中管成熟。为了验证这个假设，我们将完成以下操作 三个具体目标：目标。1确定Osi基因在顶腔运输中的功能 管成熟过程中的基质。瞄准2：确定Osi基因作为协调者的功能， 控制贩运活动的组成部分。瞄准3：鉴定直接与Osi蛋白结合的蛋白质。履行 这些目标，我们将产生携带内体标记，腔蛋白标记的果蝇品系， 在Osi突变体背景中，以确定Osi中的内体和腔蛋白的变化 变种人然后，我们将产生在Osi中表达Osi转基因或内体转基因的细胞系 突变体来分析救援效果。此后，我们将产生携带个体的苍蝇菌株， Osi突变体中的内体突变以测试Osi基因和内体之间的遗传相互作用 基因.最后，我们将在气管中表达HA标记的Osi基因，以分离相互作用的蛋白质。 Osi蛋白质。 因此，为了进行拟议的研究，携带多种突变的蝇株， 平衡器和转基因已经产生并将产生。目前， 在实验室室温下。不幸的是，该项目产生的几十种菌株 由于温度的意外显著降低或升高而损失。拟议 仪器，飞室，型号DR-36 VL，从日内瓦科学，保持适当的 湿度、光照周期和稳定的温度将使果蝇品种不仅存活，而且茁壮成长。 没有它，就不可能及时进行拟议的实验。

This proposal is for the acquisition of a Drosophila chamber to replace an old and broken chamber, which was used to culture Drosophila strains in our laboratory. The awarded NIH R15 research project is to study the role of a poorly understood Osiris (Osi) gene family in regulating tube maturation in Drosophila trachea, which is a premier system to reveal fundamental mechanisms of tubular organ formation. Malformation of tubes leads to various human diseases, such as polycystic kidney disease and vascular diseases. The Drosophila trachea is a ramifying network of epithelial tubes with a monolayer of epithelial cells surrounding an apical lumen. During tube maturation, the apical secretion burst deposits large amounts of luminal matrix components to the apical extracellular lumen. Then, this lumen is cleared before air can fill the lumen. Previous studies have shown that components of protein trafficking pathways have been implicated in the secretion/clearance of luminal proteins. The objective of the awarded project is to reveal the functions of Osi proteins as “broader coordinators” of protein trafficking during tube maturation. To test this hypothesis, we will complete the following three specific aims: Aim. 1 Determine the function of Osi genes in trafficking of apical luminal matrix during tube maturation. Aim. 2: Determine the function of Osi genes as coordinators to control trafficking components. Aim. 3: Identify proteins that directly bind to Osi proteins. To fulfill these goals, we will generate fly strains that carrying endosome markers, luminal protein markers in Osi mutant background to determine changes of endosomes and luminal proteins in Osi mutants. Then, we will generate lines to express Osi transgenes or endosome transgenes in Osi mutants to analyze the rescue effect. Thereafter, we will generate fly strains carrying individual endosome mutations in Osi mutants to test genetic interaction between Osi genes and endosome genes. Finally, we will express HA-tagged Osi genes in trachea to isolate proteins that interact with Osi proteins. Therefore, to carry out the proposed research, fly strains carrying multiple mutations, balancers, and transgenes have been and will be generated. Currently the fly strains are cultured at room temperature in the lab. Unfortunately dozens of strains that were generated for the project were lost due to unexpected significant decrease or increase of the temperatures. The proposed instrument, a fly chamber, Model DR-36VL, from GENEVA SCIENTIFIC maintains proper humidity, light cycles, and stable temperatures will keep the fly strains not only alive but thriving. Without it, it will not be possible to carry out the proposed experiments in a timely manner.