Using the C. elegans Oocyte to Model the Cell Biology of Early Onset Dystonia

使用线虫卵母细胞模拟早发性肌张力障碍的细胞生物学

• 批准号: 9021284
• 负责人: David Irwin Greenstein
• 金额: $ 22.8万
• 依托单位: UNIVERSITY OF MINNESOTA
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-08-15 至 2017-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9021284
• 关键词: Address; Age; Animals; Biochemical; Biological; Caenorhabditis elegans; Cell Nucleus; Cell physiology; Cells; Cellular biology; Child; Defect; Development; Dystonia; Early Onset Dystonia; Embryo; Embryonic Development; Essential Tremor; Experimental Models; Genes; Genetic; Genetic Models; Genetic Screening; Glutamic Acid; Goals; Growth; Homologous Gene; Human; Image; Inherited; Life; Limb structure; Mediating; Meiosis; Messenger RNA; Modeling; Molecular; Molecular Genetics; Movement Disorders; Muscle Contraction; Mutation; Nematoda; Neurologic; Neurons; Nuclear Envelope; Nuclear Export; Oocytes; Oogenesis; Parkinson Disease; Pathogenesis; Pathway interactions; Positioning Attribute; Posture; Prevalence; Process; Protein Biosynthesis; Proteins; Recovery of Function; Research; Resolution; Ribonucleoproteins; Role; Site; Suppressor Mutations; Symptoms; Synapses; System; Testing; TorsinA; Translating; Translational Regulation; Walking; Work; base; cellular imaging; early onset; experience; functional restoration; genetic manipulation; high reward; high risk; insight; mRNA Export; mutant; neuromuscular function; novel; particle; protein complex; protein function; restoration; temporal measurement; tool; trafficking

ABSTRACT Early onset dystonia or DYT1 dystonia is a debilitating neurological movement disorder that first presents in children at a mean age of about 12. DYT1 dystonia is caused by a mutation in the DYT1/Tor1a gene that encodes the evolutionarily conserved torsinA protein resulting in the deletion of a single glutamic acid residue (ΔE302/303 or ΔE). The mechanism through which the ΔE mutation causes DYT1 dystonia is unclear because the basic cellular function of torsinA is unknown. Intriguing new work suggests that torsinA might function in a new cellular mechanism by which large ribonucleoprotein particles (RNPs) are exported from the nucleus via budding through the nuclear envelope. When this function of torsinA is perturbed, messenger RNAs appear to inefficiently traffic to their synaptic sites of protein synthesis, compromising neuromuscular function. The field needs to know whether this new model is correct, and if so, to identify the specific molecular mechanisms by which torsinA promotes nuclear envelope budding for RNP export. This application seeks to address these goals using the oocytes of the nematode Caenorhabditis elegans as a tractable experimental model. Mutations in the best characterized C. elegans torsinA homolog (called ooc-5 for oocyte formation abnormal five) cause defects in oocyte growth. The oocyte growth defect is the earliest developmental abnormality observed in ooc-5 mutants and therefore is likely reflective of the primary underlying biochemical and cell biological deficits observed in cells lacking torsinA/OOC-5 function. Our hypothesis is that ooc-5 mutations disrupt oogenesis by interfering in part with the nuclear export assembly, or function of oocyte growth-promoting RNPs that we have defined. Given its extensive evolutionary conservation, OOC-5/torsinA is likely to perform the same elemental protein function in C. elegans oocytes and mammalian neurons. Interestingly, prior work has shown that the mechanisms of translational regulation discovered in C. elegans oocytes are also important for the development and function mammalian neurons. This proposal capitalizes on the many experimental advantages afforded by the C. elegans germline system, including the ability to conduct high-resolution live-cell imaging and the ease of molecular genetic and biochemical manipulations. Further, our group has spent two decades pioneering the mechanisms controlling oocyte development in C. elegans. Recently, we defined proteins and mRNA components of RNPs that regulate the growth, meiotic development, and developmental potential of C. elegans oocytes. Our research team is thus ideally positioned to critically test the generality of an RNP budding role for torsinA. To assess this new role for torsinA, we will: (1) Analyze the cellular mechanisms by which OOC-5 controls the oocyte growth process; and (2) Define genetic networks that can restore function to ooc-5/torsinA mutants.

抽象的 早发性肌张力障碍或 DYT1 肌张力障碍是一种使人衰弱的神经运动障碍，首先出现在 平均年龄约为 12 岁的儿童。DYT1 肌张力障碍是由 DYT1/Tor1a 基因突变引起的， 编码进化上保守的torsinA蛋白，导致单个谷氨酸残基的缺失 （ΔE302/303 或 ΔE） ΔE 突变导致 DYT1 肌张力障碍的机制尚不清楚，因为 TorsinA 的基本细胞功能尚不清楚，有趣的新研究表明，torsinA 可能在细胞中发挥作用。 大核糖核蛋白颗粒（RNP）通过新的细胞机制从细胞核中输出 当torsinA的这种功能受到干扰时，信使RNA似乎会从核膜中出芽。 无法有效地运输到蛋白质合成的突触位点，从而损害神经肌肉功能。 需要知道这个新模型是否正确，如果正确，则通过以下方法确定具体的分子机制 其中torsinA促进RNP输出的核膜出芽。 本申请旨在利用线虫的卵母细胞来实现这些目标 线虫作为易于处理的实验模型。 线虫 torsinA 同源物的突变。 （卵母细胞形成异常五称为ooc-5）导致卵母细胞生长缺陷卵母细胞生长缺陷。 在 ooc-5 突变体中观察到的最早的发育异常，因此可能反映了 在缺乏torsinA/OOC-5功能的细胞中观察到的主要潜在生化和细胞生物学缺陷。 我们的假设是 ooc-5 突变通过部分干扰核输出来破坏卵子发生 鉴于其广泛的进化，我们已经定义了促进卵母细胞生长的 RNP 的组装或功能。 保守性，OOC-5/torsinA 可能在秀丽隐杆线虫卵母细胞中发挥相同的基本蛋白功能 先前的工作已经表明哺乳动物神经元的翻译调节机制。 在秀丽隐杆线虫卵母细胞中发现的对哺乳动物神经元的发育和功能也很重要。 该提案利用了线虫种系系统提供的许多实验优势， 包括进行高分辨率活细胞成像的能力以及分子遗传学和 此外，我们的团队花了二十年时间探索控制机制。 最近，我们定义了 RNP 的蛋白质和 mRNA 成分。 调节线虫卵母细胞的生长、减数分裂发育和发育潜力。 因此，团队非常适合严格测试 TorsinA 的 RNP 萌芽作用的普遍性，以评估这一点。 对于torsinA的新作用，我们将：（1）分析OOC-5控制卵母细胞生长的细胞机制 过程；和（2）定义可以恢复 ooc-5/torsinA 突变体功能的遗传网络。