Characterization of genes required for border cell migration in Drosophila

果蝇边界细胞迁移所需基因的表征

• 批准号: 8866193
• 负责人: Lauren M. Anllo
• 金额: $ 4.31万
• 依托单位: PRINCETON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-07-01 至 2016-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8866193
• 关键词: Abnormal Cell; Actins; Address; Adhesions; Affect; Animals; Anterior; Antibodies; Architecture; Behavior; Behavioral; Biological Assay; Biological Models; Cancer Biology; Cell Adhesion; Cell Culture Techniques; Cell Polarity; Cells; Complex; Culture Media; Cyclic AMP; Cyclic AMP-Dependent Protein Kinases; Defect; Destinations; Development; Disease; Drosophila genus; E-Cadherin; Epithelial Cells; Epithelium; Exhibits; Fertility; Follicle Stimulating Hormone; G-Protein-Coupled Receptors; Gene Expression; Genes; Genomics; Health; Homologous Gene; Human; Human Development; Image; Immigration; LH Receptors; Life; Ligands; Link; Location; Malignant Neoplasms; Mammalian Cell; Maps; Membrane; Migration Assay; Modeling; Molecular; Morphology; Motion; Mutation; Neoplasm Metastasis; Nervous system structure; Nurses; Oocytes; Oogenesis; Organ; Organism; Ovary; Pathway interactions; Pattern; Phenotype; Play; Process; Proteins; RNA Interference; Regulation; Reporter; Rickets; Role; Running; Signal Transduction; Speed; System; Testing; Time; Traction; Wing; Work; Wound Healing; analog; beta catenin; bursicon; cancer cell; cell motility; egg; epithelial to mesenchymal transition; experience; human disease; insight; malignant breast neoplasm; migration; monolayer; mutant; nervous system development; novel; prevent; receptor; research study; stem cell niche; success; tissue culture; wound

DESCRIPTION (provided by applicant): During normal development of an organism, cells migrate from their original location to form specific organs, to populate stem cell niches and to form the complex nervous system. Abnormal cell migration is also observed in invasive cancer situations. In order to reveal the underlying mechanisms of cell migration, model systems such as the Drosophila ovary have been important in the past. Nevertheless, many questions about the regulation of cell migration are still unresolved, and this proposal addresses the role of a novel factor, Rickets, that we have found to be important for normal border cell migration in the Drosophila egg chamber. Rickets encodes a G-protein coupled receptor, and we have observed that border cells mutant for rk often fail to reach their destination at the anterior end of the oocyte. This proposal aims to investigate the underlying molecular causes of the migration defects and to understand the pathway in which Rk acts to promote cell migration. In Aim 1, I will identify the cellular architecture that is altered in the mutant cells. This will involve testng the level and distribution of important cell adhesion regulators, such as E-cadherin and Beta-Catenin, and polarity markers such as PAR-1 and PAR-3 in the mutant cells. I will also test whether Rk affects JAK/STAT signaling. In Aim 2, I will perform live imaging approaches to determine precisely what aspect of cell migration is altered in the mutant cells. I will focus on te transition of the border cells from epithelial to migratory morphology, on the detachment of the cells from the epithelium, on the speed and actual motion of the cells during migration (tumbling versus running) and on the extension of actin rich protrusions from the migrating cells. I will als test whether PKA and cAMP are acting downstream of Rk in the border cells. In Aim 3, I will test whether the effects or Rk on migration can also be observed in mammalian tissue culture. For this I will use an established system of breast cancer-derived cells that can be manipulated by RNAi approaches. A previously described scratch assay will allow me to determine whether mammalian cells with knockdown levels of Rk exhibit the same defects in migration as the mutant border cells in Drosophila. The proposed work will therefore define a new input into cell migration in Drosophila, and provide novel insights into this important developmental mechanism that are relevant to human development and disease.

描述（由申请人提供）：在生物体的正常发育过程中，细胞从其原始位置迁移到形成特定器官，填充干细胞龛并形成复杂的神经系统。在浸润性癌症中也观察到异常的细胞迁移。为了揭示细胞迁移的潜在机制，像果蝇卵巢这样的模型系统在过去一直很重要。尽管如此，关于细胞迁移调控的许多问题仍未解决，该建议解决了一个新因素佝偻病的作用，我们发现佝偻病对果蝇卵腔中正常边界细胞迁移很重要。佝偻病编码一种g蛋白偶联受体，我们观察到rk突变的边界细胞经常不能到达卵母细胞前端的目的地。本研究旨在研究迁移缺陷的潜在分子原因，并了解Rk促进细胞迁移的途径。在目标1中，我将确定突变细胞中发生改变的细胞结构。这将包括测试突变细胞中重要的细胞粘附调节因子的水平和分布，如E-cadherin和β - catenin，以及极性标记物，如PAR-1和PAR-3。我还将测试Rk是否影响JAK/STAT信令。在目标2中，我将执行实时成像方法，以准确确定突变细胞中细胞迁移的哪个方面发生了改变。我将重点关注边界细胞从上皮细胞到迁移形态的转变，细胞从上皮细胞的脱离，细胞在迁移过程中的速度和实际运动（翻滚与奔跑），以及迁移细胞中富含肌动蛋白的突起的延伸。我还将测试PKA和cAMP是否在边界细胞中作用于Rk的下游。在Aim 3中，我将测试Rk对迁移的影响是否也可以在哺乳动物组织培养中观察到。为此，我将使用一个可以通过RNAi方法操纵的乳腺癌来源细胞的既定系统。先前描述的划痕试验将允许我确定具有Rk敲低水平的哺乳动物细胞是否表现出与果蝇突变边界细胞相同的迁移缺陷。因此，提出的工作将为果蝇细胞迁移提供新的输入，并为这一与人类发育和疾病相关的重要发育机制提供新的见解。