CELLULAR ROLE OF GRANINS IN HORMONE AND NEUROTRANSMITTER STORAGE AND R

谷粒蛋白在激素和神经递质储存和 R 中的细胞作用

基本信息

批准号：
7957615
负责人：
LAURENT TAUPENOT
金额：
$ 0.78万
依托单位：
UNIVERSITY OF CALIFORNIA, SAN DIEGO
依托单位国家：
美国
项目类别：
财政年份：
2009
资助国家：
美国
起止时间：
2009-06-15 至 2010-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7957615
关键词：
Adrenal Medulla Biogenesis C-terminal Cells Chimera organism Chromaffin Cells Chromaffin granule Chromogranin A Chromogranins Computer Retrieval of Information on Scientific Projects Database Endocrine Exocytosis Family Funding Grant Hormones Image Analysis Institution Luminescent Proteins Mediating Membrane Molecular Mus Neuroendocrine Cell Neurons Neurosecretory Systems Neurotransmitters Organelles PHluorin Pathway interactions Peptide Fragments Protein Region Proteins Proteolytic Processing Recruitment Activity Reporting Research Research Personnel Resources Role Secretory Vesicles Series Sorting - Cell Movement Source Structure Sympathetic Nervous System Testing Total Internal Reflection Fluorescent United States National Institutes of Health Variant chromogranin B chromogranin C novel response secretogranins secretory protein trafficking vacuolar H+-ATPase

项目摘要

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. The chromogranins/secretogranins (granins) are a family of regulated secretory proteins widely distributed in dense-core secretory granules (DCGs) of neuroendocrine, endocrine, and neuronal cells, and includes chromogranin A (CgA), chromogranin B (CgB), and secretogranin II (SgII). Granins function as pro-hormones, giving rise by proteolytic processing to biologically active peptide fragments. Recent evidence suggests that granins may be required for the biogenesis of DCGs, and hormone sequestration in neuroendocrine cells. Using a series of novel granin chimeric photoproteins, we aim to provide a molecular definition of the cellular role of granins in hormone and neurotransmitter storage and release within the sympathetic nervous system, and the adrenal medulla. This project aims to characterize the contribution of CgA to the initiation and formation of DCGs in sympathoadrenal chromaffin cells, and to identify putative granulogenic determinant(s) within the primary structure of this granin. We previously reported that information necessary for CgA trafficking within the regulated pathway of chromaffin cells is contained in the amino-terminal region (CgA1-115) of the granin1. More recently, we have shown that a V-ATPase-mediated acidification of the TGN and/or the immature chromaffin granule contributes to the sorting of CgA to the regulated secretory pathway2 and to the biogenesis of DCGs by a mechanism that may recruit a sorting/granulogenic determinant located within CgA1-115, but not the C-terminal region of the protein 2, 3 We will test the hypothesis that CgA may rescue the formation of secretory granules in secretory organelle-deficient chromaffin cells, and a particular motif (cis determinant) within the primary structure of CgA is essential for such granulogenic role. We propose to express a series of CgA domains fused to EGFP or pHluorin in secretory organelle-deficient chromaffin cells, to investigate and characterize granulogenic cis determinant(s) within the primary structure of CgA. Responses to be investigated include secretagogue-stimulated exocytosis of transiently transfected CgA chimera by TIRF microscopy. The studies will be initially conducted in the sympathoadrenal cell variant A35C, which lacks DCGs and a regulated secretory pathway, and does not express several membrane or soluble secretory granule proteins, including CgA, CgB, and SgII 4. We will extend these studies to primary culture of chromaffin cells isolated from CgA-/- KO mice, which show reduced DCGs number.

该副本是利用众多研究子项目之一由NIH/NCRR资助的中心赠款提供的资源。子弹和调查员（PI）可能已经从其他NIH来源获得了主要资金，因此可以在其他清晰的条目中代表。列出的机构是对于中心，这不一定是调查员的机构。铬甘氨酸蛋白/分泌蛋白（花格替植物）是神经内分泌，内分泌和内分泌细胞的密集核分泌颗粒（DCGS）中广泛分布的家族，包括铬烷蛋白A（CGA），CGA），CHOMOGRANIN B（CGGB）B（CGBB（CGBB）和Socretrogranin II（Sgigogranin II）（Sgiii）（Sgiii）（Sgii）（Sgii）（Sgii）格拉蛋白起促激素的作用，通过蛋白水解加工产生生物活性肽片段。最近的证据表明，GRANIN可能需要Granins的生物发生，而神经内分泌细胞中的激素隔离。使用一系列新型的花岗岩嵌合光蛋白，我们旨在提供分子定义加拉蛋白在激素和神经递质储存中的细胞作用，并在交感神经系统中以及肾上腺髓质中释放。该项目的目的是表征CGA对交感神经膜细胞中DCG的启动和形成的贡献，并鉴定该花岗岩主要结构内的假定颗粒状决定因素。我们先前报道说，在铬蛋白细胞的调节途径中，CGA运输所需的信息包含在花格替1的氨基末端区域（CGA1-115）中。最近，我们表明，TGN和/或未成熟的铬脂颗粒的V-ATPase介导的酸化有助于将CGA分类为受调节的分泌途径2和DCG的生物发生，而DCG的生物发生通过一种机制，该机制可以通过一种可能募集CGA1-115中的分类/颗粒状确定性的机制，但不是位于CGA1-115的区域中，但是属于CGA1-115的区域的。我们将检验以下假设：CGA可以挽救分泌细胞器缺陷型铬蛋白细胞中分泌颗粒的形成，而CGA的主要结构中的特定基序（CIS依赖性）对于这种颗粒作用至关重要。我们建议在分泌细胞器缺陷型铬蛋白细胞中表达一系列与EGFP或phluorin融合的CGA结构域，以研究和表征CGA的主要结构内的颗粒状成分CIS决定因素。对研究的反应包括通过TIRF显微镜对暂时转染的CGA嵌合体的促分泌刺激的胞吐作用。这些研究最初将在缺乏DCG和受调节的分泌途径的交感肾上腺细胞变体A35C中进行，并且不表达几种膜或可溶性分泌颗粒蛋白，包括CGA，CGB，CGB和SGII 4。