GANGLIOSIDE GM1-MEDIATED TRAFFICKING IN INTESTINAL CELLS

神经节苷脂 GM1 介导的肠细胞转运

• 批准号: 6227498
• 负责人: Anne Armour Wolf
• 金额: $ 12.04万
• 依托单位: BOSTON CHILDREN'S HOSPITAL
• 依托单位国家: 美国
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-07-01 至 2006-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6227498
• 关键词: Golgi apparatus; apical membrane; basolateral membrane; cellular polarity; cholera toxin; confocal scanning microscopy; endocytosis; endoplasmic reticulum; gangliosides; gastrointestinal absorption /transport; gastrointestinal epithelium; immunocytochemistry; intracellular transport; lipid structure; membrane lipids; tissue /cell culture; transcytosis

DESCRIPTION (adapted from the application) Research: The goal of this proposal is to elucidate the itinerary and mechanisms of endocytosis and intracellular trafficking mediated by ganglioside GM1 in polarized intestinal epithelia. We recently demonstrated that ganglioside GM1 drives the association of cholera toxin (CT) with Detergent Insoluble Membrane Domains (DIGs) and that this association is necessary for toxin action in T84 cells. Preliminary studies now suggest that GM1-mediated signal transduction depends on membrane cholesterol and the ceramide structure of GM1. The studies outlined in this research proposal will use CT and related toxins to further examine the mechanisms by which GM1 targets ligands into and across the intestinal epithelial cell. S12. Aim 1 will identify structural features of GM1 which determine association with DIGs and subsequent trafficking into intestinal cells. Specifically, we will examine the effects on trafficking of altering: 1) GM1 ceramide or oligosaccharides structure; 2) membrane cholesterol; and 3) ganglioside clustering. Sp. Aim 2 will demonstrate the specificity of GM1 as a targeting motif for 1) endocytosis, 2) retrograde trafficking into the Golgi and ER, and 3) transcytosis. Sp. Aim 3 will compare GM1-dependent endocytosis via apical and basolateral cell membranes. In particular, we will compare 1) how GM1-binding at apical and basolateral cell surfaces affects the efficiency of toxin uptake, and 2) whether toxin bound to apical or basolateral GM1 eventually colocalize in the apical recycling endosome. Candidate: Dr. Wolf is a Board Certified pediatric gastroenterologist and is an Instructor of Pediatrics at Children's Hospital. She has been involved in basic research since high school and is dedicated to a career in academic medicine. Her initial findings have resulted in a first-author publication in The Journal of Cell Biology. Studies for two other manuscripts are near completion. Dr. Wolf will continue to train under the mentorship of Dr. Lencer, in a supportive and productive research environment, as she transitions to independent work. Environment: Dr. Wolf will conduct her research in Dr. Wayne Lencer's laboratory at Children's Hospital. She has full 75% protected time, assigned desk and bench space, and access to all supplies and equipment necessary to complete the proposed research program. She will continue to participate in regularly scheduled lab meetings and joint seminars in cell biology. The richness of educational opportunities and individuals available to Dr. Wolf on campus at Harvard Medical School will foster her development as an independent investigator capable of determining her own future directions.

描述（改编自应用程序） 研究：该提案的目标是阐明行程和 神经节苷脂介导的内吞作用和细胞内运输机制 极化肠上皮细胞中的 GM1。我们最近证明了 神经节苷脂 GM1 驱动霍乱毒素 (CT) 与洗涤剂的关联 不溶性膜结构域 (DIG) 并且这种关联对于 T84 细胞中的毒素作用。目前初步研究表明 GM1 介导 信号转导取决于膜胆固醇和神经酰胺结构 GM1。本研究提案中概述的研究将使用 CT 和相关技术 毒素以进一步检查 GM1 靶向配体进入和 穿过肠上皮细胞。 S12。目标 1 将确定结构 GM1 的特征决定了与 DIG 和后续的关联 贩运到肠细胞中。具体来说，我们将研究对 改变的运输：1) GM1神经酰胺或寡糖结构； 2） 膜胆固醇； 3）神经节苷脂聚类。 Sp。目标 2 将展示 GM1 作为 1) 内吞作用、2) 逆行靶向基序的特异性 转运至高尔基体和内质网，以及 3) 转胞吞作用。 Sp。目标3将进行比较 通过顶端和基底外侧细胞膜进行 GM1 依赖性内吞作用。在 特别是，我们将比较 1) GM1 在顶端细胞和基底外侧细胞的结合情况 表面影响毒素吸收的效率，2）毒素是否结合 顶端或基底外侧 GM1 最终在顶端回收中共定位 内体。 候选人：Wolf 博士是一名委员会认证的儿科胃肠病学家，也是一名 儿童医院儿科讲师。她曾参与基础 从高中起就开始研究，并致力于学术医学事业。 她的初步发现已在《杂志》上以第一作者发表 细胞生物学。另外两份手稿的研究已接近完成。博士。 沃尔夫将继续在伦瑟博士的指导下进行训练，并提供支持 当她过渡到独立工作时，她拥有富有成效的研究环境。 环境： Wolf 博士将在 Wayne Lencer 博士的实验室进行研究 儿童医院实验室。她有 75% 的受保护时间，已分配 办公桌和长凳空间，以及获得所有必要的用品和设备 完成拟议的研究计划。她将继续参加 定期安排细胞生物学实验室会议和联合研讨会。这 沃尔夫博士可以获得丰富的教育机会和个人 哈佛医学院的校园将促进她作为独立的人的发展 调查员能够确定自己未来的方向。