Exploring how cells generate and release distinct subpopulations of dense-core vesicles

探索细胞如何产生和释放不同的致密核心囊泡亚群

• 批准号: 10679873
• 负责人: Michael Ailion
• 金额: $ 23.33万
• 依托单位: UNIVERSITY OF WASHINGTON
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-04-15 至 2025-03-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10679873
• 关键词: Address; Afferent Neurons; Behavior; Behavior Control; Behavior Disorders; Beta Cell; Biogenesis; Biogenic Amines; Biological; Biological Models; Caenorhabditis elegans; Calcium; Cell Line; Cells; Cellular biology; Characteristics; Data; Defect; Dense Core Vesicle; Disease; Dopamine; Electron Microscopy; Endocrine; Endosomes; Family; Functional disorder; Future; Generations; Heterogeneity; Individual; Insulin; Investigation; Knowledge; Lipids; Mental disorders; Metabolic Diseases; Metabolism; Molecular; Mood Disorders; Nematoda; Nerve Growth Factors; Neurologic; Neuromodulator; Neurons; Neuropeptides; Norepinephrine; Organelles; Pathway interactions; Pattern; Peptides; Physiological; Physiology; Population; Process; Protein Isoforms; Proteins; Rattus; Recycling; Role; Sensory; Signaling Molecule; Site; Sorting; Structure of beta Cell of islet; System; Testing; Vesicle; cell type; experimental study; insight; member; monoamine; nervous system disorder; peptide hormone; protein complex; rab GTP-Binding Proteins; segregation; sensor; synaptotagmin; vesicular release

Project summary When examined by electron microscopy, neurons can be seen to carry organelles that look like little black dots. These black dots are called dense-core vesicles and they carry many important transmitters that act as neuromodulators, including neuropeptides, nerve growth factors, and monoamines such as dopamine and norepinephrine. Such dense-core vesicle cargos regulate a wide array of behaviors, and defects in such cargos can contribute to numerous mood disorders and other neurological conditions. However, little is understood about the cell biology of how dense-core vesicles are made, acquire cargos and mature, are trafficked to release sites, and ultimately released. Thus, the little black dots in neurons are really a big black box. Adding to the mysteries and complexity of these organelles, many neurons carry multiple dense-core vesicle cargos in the same cell. Are these different cargos copackaged together in the same dense-core vesicles or are they packaged separately in distinct vesicles? The answer to this question is surprisingly known in only a few cases, and the general pattern of copackaging versus segregation of distinct cargos is unclear, but is of key physiological relevance as it determines whether different dense-core vesicle cargos are coreleased or can be released independently. Additionally, in cases where distinct dense-core vesicle subpopulations are known to exist in the same cell, it is unclear how these distinct populations are generated and how cargos are differentially sorted. To begin to address these gaps in understanding, here we aim to establish two model systems for the study of distinct dense-core vesicle populations in the same cell: the ASI sensory neuron in the nematode C. elegans and the rat pancreatic beta-cell line 832/13. Our preliminary data show that members of a known dense-core vesicle biogenesis and maturation pathway, the Rab2/EARP pathway, are required for one subpopulation of dense-core vesicles in both the ASI neuron and the 832/13 cell line, but not for another subpopulation in the same cell. In Aim 1, we will further define the requirements for members of the Rab2/EARP pathway in the biogenesis and maturation of distinct dense-core vesicle subpopulations in the ASI neuron and 832/13 cells. Additionally, we will perform candidate screens in an attempt to identify factors required for the Rab2/EARP-independent generation of dense-core vesicles in both cell types. In Aim 2, we will test the hypothesis that different subpopulations of dense-core vesicles are marked by different isoforms of the synaptotagmin family of calcium sensors. We will also determine whether these synaptotagmins control the release of distinct dense-core vesicle subpopulations. In summary, this project will identify the basic molecules required for the generation and release of distinct subpopulations of dense-core vesicles in the same cell, and set the stage for more mechanistic investigations into how these processes occur.

项目总结 当用电子显微镜检查时，可以看到神经元携带看起来像小黑点的细胞器。 这些黑点被称为致密核小泡，它们携带着许多重要的递质，这些递质充当 神经调节剂，包括神经肽、神经生长因子和单胺类物质，如多巴胺和 去甲肾上腺素。这种致密核心囊泡货物调节广泛的一系列行为，并且在这样的 货物可能会导致许多情绪障碍和其他神经疾病。然而，几乎没有什么是 了解致密核心囊泡是如何形成、获得货物和成熟的细胞生物学， 被贩卖到释放地点，并最终被释放。因此，神经元中的小黑点实际上是一个大黑 盒。使这些细胞器更加神秘和复杂的是，许多神经元携带多个致密核心 同一细胞中的囊泡货物。这些不同的货物是不是一起装在同一个致密的核心里？ 囊泡还是分开包装在不同的囊泡中？令人惊讶的是，这个问题的答案是已知的 仅在少数情况下，不同货物的共同包装和分离的一般模式尚不清楚， 但它具有关键的生理相关性，因为它决定了不同的致密核小泡货物是否 共同发布或可以独立发布。此外，在明显的致密核小泡的情况下 亚群已知存在于同一细胞中，目前尚不清楚这些不同的种群是如何产生的 以及货物如何进行差异化分类。为了开始解决这些理解上的差距，我们在这里的目标是 为研究同一细胞中不同的致密核小泡群体建立两个模型系统：ASI 线虫和大鼠胰腺β细胞系832/13中的感觉神经元：我们的初步数据 显示了已知的致密核心囊泡生物发生和成熟途径的成员，Rab2/Earp 在ASI神经元和832/13细胞中，致密核小泡的一个亚群都需要 行，但不是针对同一细胞中的另一个亚群。在目标1中，我们将进一步定义以下要求 Rab2/Earp通路成员在致密核小泡生物发生和成熟中的作用 ASI神经元和832/13细胞的亚群。此外，我们将在一个 试图确定在两者中非依赖Rab2/Earp产生致密核小泡所需的因素 单元类型。在目标2中，我们将检验这样一种假设，即不同亚群的致密核小泡被标记 通过不同亚型的突触素家族钙传感器。我们还将确定这些 突触聚集蛋白控制着不同致密核小泡亚群的释放。总而言之，这个项目将 确定产生和释放不同的致密核心亚群所需的基本分子 并为更机械地研究这些过程是如何进行的奠定了基础 发生。