Establishment of Dpp Activity Gradient in Drosophila

果蝇 Dpp 活性梯度的建立

基本信息

批准号：
7230511
负责人：
EDWIN L FERGUSON
金额：
$ 31.58万
依托单位：
UNIVERSITY OF CHICAGO
依托单位国家：
美国
项目类别：
财政年份：
1994
资助国家：
美国
起止时间：
1994-05-01 至 2009-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7230511
关键词：
Address Affect Alleles Appearance Arthropods Back Binding Biological Assay Blastoderm Bone Morphogenetic Proteins Cell Count Cell surface Cells Cellular biology Chordata Class Cleaved cell Complex Coupling Cultured Cells Development Developmental Biology Diffusion Dorsal Dose Drosophila genus Elevation Embryo Embryonic Development Endocytosis Endopeptidases Extracellular Protein Extracellular Space Feeds Generations Genes Goals Growth Factor Imagery Individual Label Lead Ligands Localized Location Methods Modeling Mutation Pathway interactions Pattern Peptide Hydrolases Perivitelline Space Play Process Protein Family Proteins Proteolysis Range Reagent Role Series Shapes Signal Pathway Signal Transduction Staging Structure System Testing Transforming Growth Factor beta Transport Process Whole Organism base blastomere structure bone morphogenetic protein receptors extracellular gain of function mutation genetic analysis in vivo insight loss of function mutation member mutant novel prevent receptor receptor internalization research study response tool trafficking ubiquitin-protein ligase

项目摘要

DESCRIPTION (provided by applicant): In both arthropods and chordates, an activity gradient of a member of the Bone Morphogenetic Protein family patterns the embryonic dorsal-ventral axis. In Drosophila, the decapentaplegic (dpp) gene is expressed at uniform intensity over the dorsal 40% of the embryo but functions in a dose-dependent fashion to promote dorsal structures. Visualization of secreted, reinternalized Dpp protein indicates that during late blastoderm stage Dpp becomes concentrated in the dorsal 10% of cells by a process of directional diffusion through the extracellular space. Understanding this novel mechanism of directed ligand transport could offer insights into the establishment and interpretation of positional information during vertebrate development. In its most simple form, this process involves formation of a protein complex that sequesters Dpp from its receptors, free diffusion of that complex through the extracellular space, and ultimate proteolysis of the complex, allowing ligand release and signaling. The experiments in the first specific aim both seek to gain greater insights concerning the function of individual components of this transport system and address the fundamental question of the initial asymmetries that are necessary for generation of directional transport. While these extracellular proteins are critical for the establishment of pattern, phenotypic analysis of specific mutations in intracellular components of the Dpp signaling pathway strongly suggests that regulated receptor turnover, possibly based on prior signaling strength, may also contribute substantially to the establishment of the final pattern of ligand localization. In the second specific aim, reagents will be developed that specifically visualize cell surface bound or reinternalized receptors. These tools will then be used to determine whether Dpp signaling during early blastoderm feeds back to control receptor levels later during development by examination of receptor distribution in embryos lacking ligand, and in embryos mutant for downstream components of the signaling pathway. The third aim is explore the role of endocytosis in the control of Dpp signaling, with the ultimate goal of coupling an understanding of the cell biology of this process with the developmental biology of early embryonic patterning. Reagents that mark specific endocytic compartments will be used to determine the subcellular location of internalized receptor-ligand complexes. Specific steps in endocytic trafficking will be perturbed and their effects on Dpp signaling will be examined. Lastly, a recent model for endocytic control of TGF-beta signaling formulated in vertebrate cell culture will be tested in the context of the whole organism.

描述（由申请人提供）：在节肢动物和弦弦中，骨形形态发生蛋白家族模式的一个活性梯度是胚胎背腹轴。在果蝇中，脱皮术（DPP）基因在背侧40％的胚胎上以均匀的强度表达，但以剂量依赖性的方式发挥作用以促进背结构。分泌的，重新内部DPP蛋白的可视化表明，在晚期胚泡期间，DPP通过通过细胞外空间的方向扩散过程浓缩了10％的细胞。了解这种定向配体运输的新型机制可以为脊椎动物发育过程中位置信息的建立和解释提供见解。在最简单的形式中，该过程涉及形成蛋白质复合物，该蛋白质复合物从其受体中隔离DPP，该复合物通过细胞外空间的自由扩散以及对复合物的最终蛋白水解，从而使配体释放和信号传导。第一个特定目的的实验都试图获得有关该运输系统各个组件功能的更多见解，并解决了产生方向运输所必需的初始不对称的基本问题。尽管这些细胞外蛋白对于建立模式至关重要，但对DPP信号传导途径的细胞内成分的特异性突变的表型分析强烈表明，受调节的受体周转率（可能基于先前的信号强度）也可能会为最终配体定位的最终模式的建立做出重大贡献。在第二个特定目的中，将开发特定的试剂，以特异性地可视化细胞表面结合或恢复受体。然后，这些工具将用于确定在早期胚泡中的DPP信号传导是否通过检查配体缺乏配体的胚胎的受体分布以及在信号传导途径下游成分的胚胎突变体中的受体分布来回控制受体水平。第三个目的是探索内吞作用在控制DPP信号传导中的作用，其最终目的是将对该过程的细胞生物学理解与早期胚胎构图的发展生物学结合在一起。标记特定内吞区室的试剂将用于确定内部受体配体配合物的亚细胞位置。内吞交易中的特定步骤将受到干扰，并将检查其对DPP信号的影响。最后，将在整个生物体的背景下测试了在脊椎动物细胞培养中表达的TGF-β信号传导的最新模型。