The Role of Torsins at the Nuclear Envelope Using a Novel C. elegans Model

使用新型线虫模型研究 Torsins 在核膜中的作用

基本信息

批准号：
8733065
负责人：
Michael James White VanGompel
金额：
$ 5.51万
依托单位：
UNIVERSITY OF CALIFORNIA AT DAVIS
依托单位国家：
美国
项目类别：
财政年份：
2013
资助国家：
美国
起止时间：
2013-09-30 至 2015-09-29
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8733065
关键词：
ATP phosphohydrolase Address Animal Model Biological Biological Assay Biological Models Biology Caenorhabditis elegans Cell Nucleus Cell Polarity Cell physiology Cellular biology Cytoplasm DYT1 gene Data Defect Development Developmental Cell Biology Developmental Process Dextrans Discipline Disease Dyes Dystonia Early Onset Dystonia Embryo Embryonic Development Endoplasmic Reticulum Event Exclusion Family Family member Functional disorder Future Genes Goals Homologous Gene Human Image Injection of therapeutic agent Label Laboratories Lead Life Link Lipodystrophy Mammalian Cell Mediating Meiosis Membrane Methods Modeling Movement Muscle Contraction Muscular Dystrophies Mutation Nematoda Neuromuscular Diseases Neurons Neuropathy Nuclear Nuclear Envelope Nuclear Outer Membrane Nuclear Pore Nuclear Pore Complex Nuclear Pore Complex Proteins Oocytes Oogenesis Pathway interactions Patients Phenotype Positioning Attribute Premature aging syndrome Protein Binding Proteins RNA Interference Regulation Reporter Research Personnel Role Rosa Staging Tertiary Protein Structure Testing TorsinA Training career dextran disease phenotype env Gene Products insight mutant novel nucleocytoplasmic transport protein complex public health relevance research study

项目摘要

DESCRIPTION (provided by applicant): Proteins in the nuclear envelope (NE) have been shown to be important regulators of cell function and development, and disruption of these proteins leads to a range of diseases termed laminopathies. The Torsin family of AAA+ ATPases have recently been proposed to regulate NE proteins. Mutation in the human gene that encodes TorsinA, DYT1, leads to the neuromuscular disease early onset dystonia. Mammalian cells with out TorsinA function have NE structural defects, and the outer nuclear membrane KASH domain protein Nesprin-3 can interact with TorsinA. However, how TorsinA functions at the NE, and how this function relates to the disease phenotype is not known. The long term goal of this project is to understand the cell biology of Torsin function at the NE, and how Torsins regulate later development. In this proposal, I will investigate Torsin function at the NE using C. elegans as a model. Our laboratory previously identified ooc-5, a C. elegans homolog of TorsinA. Mutations in ooc-5 lead to smaller oocytes and polarity and spindle positioning defects in early embryos. Interestingly, it has been shown that depletion of nucleoporins, components of nuclear pore complexes, in C. elegans leads to an ooc-5 like phenotype, suggesting a link between Torsins and nuclear pores in the worm. Furthermore, preliminary studies have revealed defects in the localization of multiple nuclear envelope proteins in ooc-5 mutants. Using GFP reporter strains and RNAi-mediated gene depletion, I propose to determine which NE protein defects are primary, and which are secondary downstream defects. I will also determine if nuclear envelope barrier and transport functions are disrupted in ooc-5 mutants by using nuclear-localized GFP markers and dextran dye injections. This will be the first test of the functional consequences of nuclear envelope defects in a Torsin mutant model. Finally, I will test which defects at the NE cause later polarity phenotypes, providing a link between the cell biology of Torsin function and the regulation of development. Results from this study will be applicable to both human Torsin function and mechanisms underlying early onset dystonia, and also to the study of embryonic polarity establishment in C. elegans.

描述（由申请人提供）：核包膜中（NE）中的蛋白质已被证明是细胞功能和发育的重要调节剂，并且这些蛋白质的破坏会导致一系列称为椎板病的疾病。最近已经提出了AAA+ ATPases的Torsin家族来调节NE蛋白。编码Torsina，Dyt1的人类基因中的突变导致神经肌肉疾病早期发作。具有torsina功能的哺乳动物细胞具有NE结构缺陷，并且外核膜Kash结构蛋白Nesprin-3可以与Torsina相互作用。然而，未知的Torsina如何在NE处发挥作用，以及该功能与疾病表型的关系。该项目的长期目标是了解NE的Torsin功能的细胞生物学，以及Torsins如何调节后来的发展。在此提案中，我将在 NE使用秀丽隐杆线虫作为模型。我们的实验室先前鉴定出了Torsina的C.秀丽隐杆线虫同源物。 OOC-5中的突变导致较小的卵母细胞，以及早期胚胎的极性和纺锤体定位缺陷。有趣的是，已经表明，在秀丽隐杆线虫中，核孔蛋白（核孔复合物的成分）的耗竭会导致OOC-5像表型这样的OOC-5，这表明蠕虫中的Torsins和核孔之间存在联系。此外，初步研究揭示了OOC-5突变体中多个核包膜蛋白定位的缺陷。使用GFP报告基因菌株和RNAi介导的基因耗竭，我建议确定哪些NE NE蛋白缺损是主要的，哪些是次级下游缺陷。我还将通过使用核位置的GFP标记和右旋染料注射核-5突变体中的核包膜屏障和运输功能在OOC-5突变体中破坏。这将是核包膜缺陷在Torsin突变模型中的功能后果的首次测试。最后，我将测试在NE导致后来的极性表型上的缺陷，从而提供了Torsin功能的细胞生物学与发育调节之间的联系。这项研究的结果将适用于早期发作肌张力障碍的人类扭曲功能和机制，也适用于秀丽隐杆线虫中胚胎极性建立的研究。