Acetylcholine Receptors in Cell Migrationis

细胞迁移中的乙酰胆碱受体

• 批准号: 10349561
• 负责人: MIHOKO KATO
• 金额: $ 32.62万
• 依托单位: CALIFORNIA INSTITUTE OF TECHNOLOGY
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-03-01 至 2024-02-29
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10349561
• 关键词: Acetylcholine; Acetylcholinesterase Inhibitors; Affect; Agonist; Aldicarb; Animals; Anterior; Bacteria; Biological Assay; Caenorhabditis elegans; Cell Polarity; Cell membrane; Cell physiology; Cells; Cholinergic Receptors; Cues; Defect; Detection; Development; Electron Microscopy; Embryo; Epithelial; Gene Expression Profiling; Genes; Gonadal structure; Human; Ion Channel; Ligands; Link; Location; Membrane; Membrane Potentials; Mesenchymal Stem Cells; Muscarinic Acetylcholine Receptor; Muscarinics; Nerve; Nervous system structure; Neurons; Nicotinic Receptors; Organ; Organism; Output; Pathway interactions; Perception; Phenotype; Plants; Process; Protozoa; Resolution; Role; Shapes; Signal Pathway; Signal Transduction; Signaling Molecule; Smooth Muscle Myocytes; Source; Specialized Epithelial Cell; Synaptic Transmission; System; Testing; Tissues; Variant; cell motility; cholinergic neuron; experimental study; gain of function; in vivo; keratinocyte; male; migration; mutant; neurotransmission; optogenetics; paralogous gene; postnatal development; receptor; response; transcriptomics

Project Summary Acetylcholine (ACh) is an ancient signaling molecule found in many organisms including bacteria, protozoa, plants, and animals. ACh detection through nicotinic and muscarinic receptors occurs in many human organs besides the nervous system but these much less studied than its neuronal function. One of the non-synaptic functions of ACh signaling is in cell migration during normal development. For example, smooth muscle cells, epithelial keratinocytes, and mesenchymal stem cells migrate in response to ACh, but a detailed mechanistic understanding of how this signaling is utilized in migration, particularly in vivo, is not available. We have developed a unique system with which to study the role of Ach signaling in cell migration in vivo. The C. elegans linker cell (LC) is a specialized epithelial cell that leads the migration of the developing male gonad. We found through single-cell transcriptomics that the LC expresses many neuronal receptors and ion channels, leading to the hypothesis that the LC uses neuronal cues for its migration. We will investigate the role of acetylcholine signaling in the LC because we observed migration defects in mutants lacking particular muscarinic and nicotinic receptors. We will initially focus on the only muscarinic receptor expressed in the LC, GAR-3, that we found to activate changes in cell orientation. We will in parallel investigate the function of the multiple nicotinic receptors in LC migration and potential cooperation between the receptor types. The LC reverses its orientation from the posterior to anterior in response to excess ACh signaling induced by treatment with the acetylcholinesterase inhibitor aldicarb, an assay we will use to identify the downstream components of the GAR-3 pathway in the LC. Since downstream targets of the branching gar-3 signaling pathway have been identified in other contexts (e.g., synaptic transmission), we will test mutants of those genes to identify the specific downstream pathway used to modulate LC orientation. To investigate the role of ACh distribution, we will reprogram the fate of neurons in the ventral nerve cord (VNC) to create artificial anterior/posterior ACh gradients to investigate how ACh distribution and amount affects LC migration. We will collaborate to use Correlation Electron Microscopy to examine GAR- 3 localization relative to the ultrastructure of LC and surrounding tissue at high resolution. We will reuse these assays to study potential effects of six nicotinic receptor subunits.

项目摘要 乙酰胆碱（ACH）是在许多生物中发现的古老信号分子，包括细菌，原生动物，， 植物和动物。通过烟碱和毒蕈碱受体进行的ACH检测发生在许多人体器官中 除了神经系统，但研究的研究远不如其神经元功能。非突触 ACH信号的功能在正常发育过程中在细胞迁移中。例如，平滑肌细胞， 上皮角质形成细胞和间充质干细胞响应于ACH，但详细的机械 了解如何在迁移中使用该信号传导，尤其是体内的迁移。我们有 开发了一个独特的系统，可以研究ACH信号在体内细胞迁移中的作用。秀丽隐杆线虫 接头细胞（LC）是一种专门的上皮细胞，导致发育中的雄性性腺的迁移。我们发现 通过单细胞转录组学表达许多神经元受体和离子通道，导致 LC使用神经元线索的迁移假设。我们将研究乙酰胆碱的作用 LC中的信号传导，因为我们观察到缺乏特定毒蕈碱和烟碱的突变体的迁移缺陷 受体。我们最初将专注于我们发现的LC GAR-3中唯一的毒蕈碱受体 激活细胞方向的变化。我们将同时研究多个烟碱受体的功能 在LC迁移和受体类型之间的潜在合作中。 LC从 响应于乙酰胆碱酯酶治疗引起的过量ACH信号的后部， 抑制剂Aldicarb，我们将使用一种测定法识别LC中GAR-3途径的下游组件。 由于在其他情况下已经确定了分支GAR-3信号通路的下游目标（例如， 突触传递），我们将测试这些基因的突变体，以识别用于的特定下游途径 调节LC方向。为了研究ACH分布的作用，我们将重新编程神经元的命运 腹神经线（VNC）创建人工前/后ACH梯度，以研究ACH分布如何 金额影响LC迁移。我们将合作使用相关电子显微镜检查GAR- 3与LC和周围组织的超微结构相对于高分辨率的定位。我们将重复使用这些 研究六个烟碱受体亚基的潜在影响的测定。