Acquisition of a Drosophila chamber for culturing Drosophila strains

获得用于培养果蝇菌株的果蝇室

• 批准号: 10580919
• 负责人: Lan Jiang
• 金额: $ 1.9万
• 依托单位: OAKLAND UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-07-01 至 2025-06-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10580919
• 关键词: Air; Apical; Autophagosome; Award; Binding; Binding Proteins; Biological Assay; Biological Models; Blood Vessels; Cessation of life; Co-Immunoprecipitations; Cytoplasmic Vesicles; Defect; Deposition; Disease; Drosophila eye; Drosophila genus; Drug Metabolic Detoxication; Endosomes; Epithelial; Epithelial Cells; Extracellular Matrix; Gases; Gene Family; Genes; Genetic; Glycocalyx; Glycolysis; Goals; Golgi Apparatus; Human; Humidity; Image; Immunohistochemistry; Individual; Insecta; Invertebrates; Kidney; Knowledge; Laboratories; Lactoylglutathione Lyase; Lead; Life; Light; Liquid substance; Lung; Lysosomes; Mediating; Metabolic; Modeling; Morphogenesis; Mutation; Organ; Organism; Pathway interactions; Phenotype; Play; Polycystic Kidney Diseases; Property; Proteins; Pyruvaldehyde; Research; Research Project Grants; Role; Shapes; Structure; System; Temperature; Testing; Time; Trachea; Transgenes; Tube; Tubular formation; United States National Institutes of Health; Vascular Diseases; Vertebrates; Vesicle; Yeasts; body system; exosome; experimental study; extracellular; fly; gene function; genetic testing; human disease; in vivo; instrument; kidney vascular structure; malformation; member; monolayer; mutant; protein biomarkers; protein transport; secretion process; spine bone structure; trafficking

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.

该提案是为了购买一个果蝇室来替换旧的和破损的室 室，用于在我们的实验室培养果蝇菌株。荣获 NIH R15 称号。 研究项目是研究一个鲜为人知的奥西里斯（Osi）基因家族在调节中的作用 果蝇气管的管成熟，这是揭示基本原理的首要系统 管状器官形成的机制。管畸形导致各种人类疾病， 例如多囊肾病和血管疾病。 果蝇气管是由单层上皮细胞组成的上皮管的分枝网络。 管腔周围的细胞在管成熟期间，顶端分泌物突然沉积。 大量的管腔基质成分到达顶端细胞外腔，然后，该管腔。 先前的研究表明，蛋白质的成分在空气充满管腔之前就被清除了。 贩运途径与腔蛋白的分泌/清除有关。 获奖项目的目标是揭示 Osi 蛋白作为“更广泛的协调员”的功能 为了检验这一假设，我们将完成以下任务。 三个具体目标： 目标 1 确定 Osi 基因在顶端管腔运输中的功能。 目标 2：确定 Osi 基因作为协调子的功能。 目标 3：识别直接与 Osi 蛋白结合的蛋白质。 这些目标，我们将产生携带内体标记、管腔蛋白标记的果蝇品系 在 Osi 突变背景中确定 Osi 中内体和管腔蛋白的变化 然后，我们将生成表达 Osi 转基因或内体转基因的品系。 此后，我们将产生携带个体的果蝇菌株。 Osi 突变体中的内体突变，用于测试 Osi 基因和内体之间的遗传相互作用 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。 与Osi蛋白。 因此，为了开展拟议的研究，携带多种突变的果蝇品系， 平衡子和转基因已经并将产生，目前果蝇品系正在培养中。 不幸的是，该项目在室温下产生了数十种菌株。 由于温度意外显着降低或升高而损失。 来自 GENEVA SCIENTIFIC 的 DR-36VL 型飞行室仪器保持正确 湿度、光循环和稳定的温度不仅可以让苍蝇品种保持活力，而且可以繁衍生息。 没有它，就不可能及时进行所提出的实验。