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Nuclear function of L1-type CAMs in the drosophila nervous system

果蝇神经系统中 L1 型 CAM 的核功能

基本信息

批准号：
9015594
负责人：
TANJA ANGELA GODENSCHWEGE
金额：
$ 44.76万
依托单位：
FLORIDA ATLANTIC UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2015
资助国家：
美国
起止时间：
2015-09-15 至 2019-09-14
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9015594
关键词：
Adult Affect Alzheimer&apos s Disease Amino Acids Ankyrins Antibodies Area Binding C-terminal Cancer cell line Cause of Death Cell Adhesion Molecules Cell Cycle Regulation Cell Line Cell Nucleus Cell membrane Cells Cytoskeleton DNA damage checkpoint Data Defect Development Disease Drosophila genus Endosomes Epitopes Gene Expression Genomics Goals Growth Health Human Image Immigration In Vitro Integral Membrane Protein Invertebrates LIS1 protein Length Life Link Lysine Malignant Neoplasms Mediating Membrane Mono-S Mutation Natural regeneration Nerve Growth Factor Receptors Nervous system structure Neural Cell Adhesion Molecule L1 Neuraxis Neurites Neuroglia Neurons Nuclear Nuclear Import Nuclear Translocation Oxidative Stress Peptide Signal Sequences Phenotype Play Polyubiquitination Post-Translational Protein Processing Protein Isoforms Proteins Purinoceptor Radioresistance Recombinants Recycling Reporting Research Role Signal Transduction Spinal cord injury Stroke Synapses Synaptic plasticity Testing Transcriptional Activation Transmembrane Domain Tyrosine Ubiquitination Vertebrates Western Blotting Work bone morphogenetic protein receptors corpus callosum hypoplasia-retardation-adducted thumbs-spastic paraparesis-hydrocephalus syndrome fly gain of function in vivo insight malignant neurologic neoplasms migration mouse model mutant neoplastic cell nervous system development nervous system disorder neurodevelopment neuroglian neuronal cell body neuropathology neuroprotection novel presenilin public health relevance research study retrograde transport secretase spinal cord regeneration trafficking

项目摘要

DESCRIPTION (provided by applicant): L1-type cell adhesion molecules (CAM) are essential for proper nervous system development. We previously demonstrated that transsynaptic signaling of the sole invertebrate L1-type CAM neuroglian (Nrg) regulates synapse growth and stability and that its function in organizing the cytoskeleton at the synapse is conserved from flies to humans. Preliminary live imaging data reveals that in the adult Nrg is retrogradely transported from the synapse to the soma in a Lissencephaly 1 (Lis1)-dependent manner. This suggests that in the adult L1-type CAMs have a yet uncharacterized function that is distinct from its well-known developmental role at the plasma membrane. Retrogradely transported L1-type CAMs may be degraded or recycled in the soma, be part of a "signaling endosome", or be retrograde signals themselves, which translocate to the nucleus. Recently, cytosolic 28kDa and 30KDa L1CAM fragments as well as a 70kDa transmembrane domain- containing fragment were shown to translocate to the nucleus. We have evidence that in addition to fragments, full-length Nrg translocates to the nuclei of Drosophila nervous systems and that Nrg fragments are likely to carry posttranslational modifications distinct from the full-length form. Our hypothesis s that some of the Nrg that is transported from the synapse to the soma will translocate to the nucleus. To test this, we will determine if the nuclear import of Nrg is reduced in Lis1 mutants when compared to wild type background and if the full- length Nrg form and its fragments carry different posttranslational modifications. In addition to nuclear translocation, we propose in aim 1 to determine mechanisms of L1-type CAM retrograde transport. We will use live imaging to determine the type of endosomal compartments in which axonal retrograde transport of Nrg occurs and the intracellular motifs that are required for Nrg retrograde transport. Recombinant expression of the intracellular domain (ICD) of L1CAM in non-neuronal cell lines altered the expression of genes involved in migration, cell cycle control and DNA damage checkpoint responses and in vitro experiments suggest that nuclear L1CAM may have a role in migration, neurite outgrowth and cellular protection against physical damage or genomic and oxidative stress caused by diseases or environmental influences. The main goal of aim 2 is to determine if nuclear L1-type CAMs have a functional role in vivo as well. For this, we will characterize the phenotypes of a mutant that lacks the transmembrane proximal nuclear localization sequence. In addition, we will transgenically express L1-type CAM proteins that mimic nuclear full- length L1-type protein and its fragments in the nervous system of Drosophila to determine if they have distinct functions by analyzing their induced phenotypes in vivo at the single cell and the organismal levels. In summary, the in vivo characterization of the mechanisms and functions of nuclear L1-type CAM signaling in the central nervous system will impact a broad range of research areas such as neuroprotection during stroke, spinal cord regeneration, Alzheimer's disease and cancer.

描述（由申请人提供）：L1型细胞粘附分子（CAM）对正常神经系统发育至关重要。我们以前证明，唯一的无脊椎动物L1型CAM神经胶质细胞（NRG）的跨突触信号调节突触的生长和稳定性，其功能在组织细胞骨架在突触是保守的从苍蝇到人类。初步的实时成像数据显示，在成人Nrg是逆行运输从突触的索马在无脑1（Lis1）依赖的方式。这表明，在成人L1型CAM有一个尚未表征的功能，这是不同于其众所周知的发展作用，在质膜。逆行转运的L1型CAM可能在索马中降解或再循环，成为“信号内体”的一部分，或者本身是逆行信号，其易位到细胞核。最近，胞质28kDa和30KDa的L1CAM片段以及70kDa的含跨膜结构域的片段被证明易位到细胞核。我们有证据表明，除了片段，全长NRG易位到果蝇神经系统的细胞核，NRG片段很可能进行翻译后修饰不同于全长形式。我们的假设是，从突触到索马的Nrg中有一部分会易位到细胞核。为了测试这一点，我们将确定当与野生型背景相比时，Nrg的核输入在Lisl突变体中是否减少，以及全长Nrg形式及其片段是否携带不同的翻译后修饰。除了核转位，我们建议在目标1，以确定机制的L1型CAM逆行运输。我们将使用实时成像来确定Nrg轴突逆行转运发生的内体隔室类型和Nrg逆行转运所需的细胞内基序。在非神经元细胞系中重组表达L1CAM的胞内结构域（ICD）改变了参与迁移、细胞周期控制和DNA损伤检查点反应的基因的表达，体外实验表明，核L1CAM可能在迁移、神经突生长和细胞保护中起作用，以防止疾病或环境影响引起的物理损伤或基因组和氧化应激。目的2的主要目标是确定核L1型CAM是否在体内也具有功能作用。为此，我们将表征缺乏跨膜近端核定位序列的突变体的表型。此外，我们将在果蝇神经系统中转基因表达模拟核全长L1型蛋白及其片段的L1型CAM蛋白，通过在单细胞和生物体水平上分析其体内诱导的表型来确定它们是否具有不同的功能。总之，在中枢神经系统中的核L1型CAM信号传导的机制和功能的体内表征将影响广泛的研究领域，如中风期间的神经保护，脊髓再生，阿尔茨海默病和癌症。