Regulation of neural development by F-box mediated ubiquitination

F-box 介导的泛素化对神经发育的调节

• 批准号: 7991107
• 负责人: ELENA M Silva
• 金额: $ 7.68万
• 依托单位: GEORGETOWN UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-07-15 至 2012-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7991107
• 关键词: 26S proteasome; Address; Affinity Chromatography; Amphibia; Area; Binding; Binding Sites; Biochemical; Bioinformatics; Biological Assay; Cell Cycle Regulation; Cell physiology; Cells; Co-Immunoprecipitations; Communities; Computer Simulation; Connective Tissue Diseases; Databases; Defect; Development; Developmental Biology; Developmental Process; Disease; Dorsal; Dwarfism; Ectoderm; Embryo; Embryonic Development; Ensure; Epidermis; Event; Excision; Expressed Sequence Tags; F Box Domain; F-Box Proteins; Family; Future; Gastrula; Geminin; Genes; Genome; Genome Scan; Genomics; Goals; Grant; In Situ Hybridization; Indium; Individual; Investigation; Knowledge; Ligase; Limb structure; Link; Location; Maintenance; Malignant Neoplasms; Mediating; Molecular Profiling; Monitor; Names; Nervous system structure; Neural Crest; Neuraxis; Neurodegenerative Disorders; Neuroectoderm; Neuroglia; Neurons; Neurula; Pattern; Pilot Projects; Play; Post-Translational Regulation; Process; Proteins; RE1-silencing transcription factor; Recording of previous events; Regulation; Research; Research Infrastructure; Research Personnel; Role; Scaffolding Protein; Screening procedure; System; Testing; Time; Transcriptional Silencer Elements; Ubiquitin; Ubiquitination; Work; Xenopus; base; cell fate specification; craniofacial; developmental neurobiology; gastrulation; genetic regulatory protein; genome sequencing; loss of function; nerve stem cell; nervous system development; neural plate; neurodevelopment; neurogenesis; novel; programs; protein degradation; protein profiling; relating to nervous system; research study; response; skills; slug; tandem mass spectrometry; tool; transcription factor; ubiquitin ligase; ubiquitin-protein ligase; zygote

DESCRIPTION (Provided by Applicant): A change in the regulatory protein profile of a cell drives fate specification. This change is in part controlled at the transcriptional level, but as cells progress to their final fate during embryonic development, regulatory proteins must also be inactivated. One mechanism is removal by ubiquitin-mediated protein degradation which involves the attachment of a multi-ubiquitin chain to target the selected proteins to the 26S proteasome for destruction. These proteins are selected by ubiquitin ligases (E3), and while there are numerous E3 ligases, the best understood are the Cullin based ligases (SCF) which contain Skp1, the scaffold protein Cullin, and the target-binding F-box proteins. There is a rich history demonstrating a role for ubiquitin-mediated protein destruction in cell-cycle regulation, and defects in this process underlie a number of cancers. Furthermore, E3 ligase deficiencies have been linked to neural crest disorders and neurodegenerative diseases, indicating that protein degradation plays a key role throughout embryonic development. While the multistep process of ubiquitination is well understood and targets have been identified, we know less about its function in the regulation of cell fate specification in vertebrate development. To gain a better understanding of the role of protein degradation in neural development, the investigators propose to identify all F-box proteins in Xenopus and use gain- and loss-of-function assays to characterize those expressed in the nervous system throughout development and in the epidermis during neural plate specification. With the completion of these aims, they will then be able to identify F-box binding partners and therefore potential targets whose degradation is required for ectodermal fate specification as well as for progression from neural precursor to differentiated neuron. F-box proteins are the largest class of E3 substrate-interacting proteins and since many bind multiple substrates, they are likely to facilitate the ubiquitination of hundreds of proteins in a variety of cellular and developmental processes. Using bioinformatic tools, thus far the investigators have identified 58 F-box proteins in Xenopus and classified them into three groups based on their protein interaction domain. Furthermore, with in silico genome scanning for REST binding sites, they have identified three F-box proteins that are expressed in the nervous system. The investigators will confirm that they play a role in ubisquitination and that they have a role in neural development by demonstrating interaction with Skp1 of the modular Cullin-based E3 ligase, and with loss-of-function analysis, respectively. The long term goal of this research program, to determine the role of these specific degradation events in the specification and differentiation of the nervous system, is fundamental to understanding the regulation of normal and disordered neural development. NARRATIVE: To transform the fertilized egg into a perfectly formed individual, specific combinations of proteins must be expressed at the right time and place. Loss of expression or misexpression of a single protein can have catastrophic consequences including cyclopia, dwarfism, connective tissue disorders, and limb as well as craniofacial defects. This application will address how the levels of proteins are controlled by degradation to ensure ordered progression through neurogenesis and therefore the proper development of the central nervous system.

描述（由申请人提供）：细胞的调节蛋白轮廓的变化驱动命运规范。 这种变化部分受到转录水平的控制，但是随着细胞在胚胎发育过程中的最终命运的发展，调节蛋白也必须被灭活。 一种机制是通过泛素介导的蛋白质降解去除，该蛋白降解涉及多泛素链的附着，以将所选蛋白靶向26S蛋白酶体进行破坏。这些蛋白质是由泛素连接酶（E3）选择的，尽管有许多E3连接酶，但最好的理解是基于Cullin的连接酶（SCF），其中包含SKP1，脚手架蛋白Cullin和靶标F-box蛋白。 有丰富的历史表明在细胞周期调节中泛素介导的蛋白质破坏的作用，并且在此过程中的缺陷是许多癌症的基础。 此外，E3连接酶缺陷与神经rest疾病和神经退行性疾病有关，表明蛋白质降解在整个胚胎发育中起着关键作用。 虽然众所周知的泛素化过程并已经确定了目标，但我们对脊椎动物发育中细胞命运规范的调节方面的功能了解较少。 为了更好地了解蛋白质降解在神经发育中的作用，研究者建议鉴定Xenopus中的所有F-box蛋白质，并使用功能障碍和功能丧失测定，以表征整个神经系统中在神经系统中以及在神经板规范期间表皮中表达的蛋白质。 随着这些目标的完成，他们将能够鉴定出F-box结合伙伴，因此，其潜在的目标是外胚层命运规范所需的降解以及从神经前体到区分神经元的发展。 F-box蛋白是最大的E3底物相互作用蛋白，由于许多结合了多个底物，因此它们很可能促进了在各种细胞和发育过程中数百种蛋白质的泛素化。 到目前为止，研究人员使用生物信息学工具鉴定了Xenopus中的58个F-box蛋白，并根据其蛋白质相互作用域将其分为三组。 此外，在用于静止结合位点的硅基基因组扫描中，他们已经鉴定出三种在神经系统中表达的F-box蛋白。 研究人员将确认他们在泛质中发挥作用，并通过证明与基于Culllin的E3连接酶的SKP1的相互作用和功能丧失分析来在神经发育中发挥作用。 该研究计划的长期目标是确定这些特定的降解事件在神经系统的规范和区分中的作用，这对于理解正常神经发育的调节至关重要。 叙述：要将受精卵变成一个完美形成的个体，必须在正确的时间和地点表达蛋白质的特定组合。 单一蛋白质的表达或表达丧失可能会带来灾难性的后果，包括环形，矮人，结缔组织疾病和肢体以及颅面缺陷。 该应用将解决如何通过降解来控制蛋白质的水平，以确保通过神经发生，从而确保中枢神经系统的正确发展。