Mechanisms of Nuclear Migration and Anchorage

核迁移和锚定机制

• 批准号: 7334778
• 负责人: DANIEL A STARR
• 金额: $ 25.17万
• 依托单位: UNIVERSITY OF CALIFORNIA AT DAVIS
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-01-01 至 2010-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7334778
• 关键词: Alleles; Animals; Binding; Biochemical; Biochemical Genetics; Biological Assay; Caenorhabditis elegans; Cell Nucleus; Cloning; Cytoplasm; Cytoskeleton; Defect; Development; Disease; Electron Microscopy; Endopeptidases; Endoplasmic Reticulum; Engineering; Enhancers; Epitopes; Eukaryota; Eukaryotic Cell; Event; Genetic; Genetic Techniques; Genome; Goals; Gold; Homologous Gene; Human; Human Development; Immigration; Immune; In Vitro; Label; Lead; Link; Localized; Location; Membrane; Microfilaments; Microtubules; Modeling; Molecular; Molecular Genetics; Muscular Dystrophies; N-terminal; Neoplasm Metastasis; Neuromuscular Junction; Normal Cell; Nuclear; Nuclear Envelope; Nuclear Inner Membrane; Nuclear Matrix; Nuclear Outer Membrane; Organelles; Pathway interactions; Peptide Hydrolases; Play; Positioning Attribute; Proteins; RNA Interference; Recruitment Activity; Role; Site; System; Temperature; Testing; Two-Hybrid System Techniques; Work; cell motility; cell type; env Gene Products; in vivo; insight; migration; nervous system disorder; novel; null mutation; polarized cell; protein function; research study

The position of the nucleus is carefully controlled in a wide variety of cell types. Nuclear migration plays a role in normal cell migration events and metastasis; defects in nuclear migration lead to the neurological disease Lissencephaly. Nuclear anchorage functions in the development of the neuro-muscular junction and may contribute to muscular dystrophy. A group of three conserved nuclear envelope proteins are required for proper nuclear positioning in C. elegans. Our objective is to characterize how these proteins function to control nuclear positioning and to identify other proteins that function with or in parallel to them. Our central hypothesis is that UNC-84 functions at the inner nuclear membrane to recruit UNC-83 and ANC-1 to the outer nuclear membrane. Together, they bridge the nuclear envelope to connect the nuclear matrix to the cytoskeleton. Our first aim will determine the topology of these three proteins using an in vivo protease protection assay and immuno-EM. Aim 2 will use molecular genetic techniques to test the central link of our model, the interaction between the SUN domain of UNC-84 and the KASH domains of UNC-83 and ANC-1. In aim 3 we expect to link UNC-83 to the cytoskeleton by identifying interacting partners through biochemical and molecular screens using essential portions of the novel domain of UNC-83. In aim 4 we take a genetic approach to identify additional proteins involved in nuclear positioning by cloning existing enhancer of unc-83 or unc-84 alleles. We will use genome-wide RNAi to screen for more enhancers. The ability to combine genetic, biochemical, and molecular approaches in a developmental system makes C, elegans a powerful system for these studies. Together, these studies will provide mechanistic insight into the fundamental problem of how the nucleus positions itself in the cytoplasm.

在多种细胞类型中，细胞核的位置受到仔细控制。 核迁移在正常细胞迁移事件和转移中发挥作用；的缺陷 核迁移导致神经系统疾病无脑畸形。核锚地 在神经肌肉接头的发育中发挥作用，并可能有助于 肌肉萎缩症。需要一组三个保守的核膜蛋白 线虫中正确的核定位。我们的目标是描述这些 蛋白质的功能是控制核定位并识别其他起作用的蛋白质 与它们一起或平行。我们的中心假设是 UNC-84 在内部发挥作用 核膜将 UNC-83 和 ANC-1 招募到外核膜。一起， 它们桥接核膜，将核基质连接到细胞骨架。我们的第一个 目标将使用体内蛋白酶保护确定这三种蛋白质的拓扑结构 测定和免疫电镜。目标2将利用分子遗传学技术来测试中心环节 我们的模型中，UNC-84 的 SUN 域和 KASH 域之间的相互作用 UNC-83 和 ANC-1。在目标 3 中，我们希望通过识别来将 UNC-83 与细胞骨架联系起来 使用生物化学和分子筛选的重要部分与合作伙伴互动 UNC-83 的新结构域。在目标 4 中，我们采用遗传方法来识别其他 通过克隆 unc-83 或 unc-84 的现有增强子参与核定位的蛋白质 等位基因。我们将使用全基因组 RNAi 来筛选更多的增强子。有能力 将遗传、生化和分子方法结合到一个发育系统中 使线虫成为这些研究的强大系统。这些研究将共同​​提供 对原子核如何将自身定位这一基本问题的机械见解 细胞质。