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.

项目概要 乙酰胆碱 (ACh) 是一种古老的信号分子，存在于许多生物体中，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乙酰胆碱检测存在于许多人体器官中 除了神经系统之外，但对这些的研究比其神经元功能要少得多。非突触之一 ACh 信号传导的功能是在正常发育过程中的细胞迁移中。例如，平滑肌细胞、 上皮角质形成细胞和间充质干细胞响应ACh而迁移，但详细的机制 目前还无法了解这种信号传导如何在迁移中，特别是在体内被利用。我们有 开发了一种独特的系统，用于研究 Ach 信号传导在体内细胞迁移中的作用。线虫 连接细胞（LC）是一种特殊的上皮细胞，引导发育中的男性性腺的迁移。我们发现 通过单细胞转录组学发现 LC 表达许多神经元受体和离子通道，从而导致 LC 使用神经元线索进行迁移的假设。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存在迁移缺陷 受体。我们首先将关注在 LC 中表达的唯一毒蕈碱受体 GAR-3，我们发现它 激活细胞方向的变化。我们将同时研究多种烟碱受体的功能 LC 迁移和受体类型之间的潜在合作。 LC 的方向与 乙酰胆碱酯酶治疗诱导的过量 ACh 信号传导从后到前 抑制剂涕灭威，我们将使用该检测来识别 LC 中 GAR-3 途径的下游成分。 由于分支 gar-3 信号通路的下游靶标已在其他情况下被识别（例如， 突触传递），我们将测试这些基因的突变体，以确定用于突触传递的特定下游途径 调节 LC 方向。为了研究乙酰胆碱分布的作用，我们将重新编程神经元的命运 腹神经索 (VNC) 创建人工前/后 ACh 梯度以研究 ACh 分布 和量影响LC迁移。我们将合作使用相关电子显微镜来检查 GAR- 3 相对于 LC 和周围组织超微结构的高分辨率定位。我们将重复使用这些 研究六种烟碱受体亚基的潜在影响的测定。