Vesicular trafficking mechanisms regulating granulocyte function

调节粒细胞功能的囊泡运输机制

• 批准号: 8703748
• 负责人: Sergio Daniel Catz
• 金额: $ 46.43万
• 依托单位: SCRIPPS RESEARCH INSTITUTE, THE
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-12-03 至 2017-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8703748
• 关键词: Actins; Adhesions; Attenuated; Azurophilic Granule; Binding; Blood; Blood Vessels; Cardiovascular Diseases; Cell membrane; Cell physiology; Chemotaxis; Communicable Diseases; Cytoplasmic Granules; Cytoskeleton; Data; Defect; Development; Disease; Endothelium; Endotoxemia; Endotoxins; Event; Exocytosis; Gelatinases; Guanosine Triphosphate Phosphohydrolases; Host Defense; Human; In Vitro; Infection; Inflammation; Inflammatory; Invaded; Laboratories; Lead; Leukocytes; Lipopolysaccharides; Mediating; Membrane; Methods; Molecular; Monomeric GTP-Binding Proteins; Movement; Mus; Organelles; Pathway interactions; Phagocytosis; Phagosomes; Physiological; Physiology; Play; Poison; Positioning Attribute; Prevention; Probability; Process; Production; Proteins; Regulation; Reperfusion Injury; Research; Role; Secretory Vesicles; Sepsis; Specificity; Systems Biology; Testing; Therapeutic; Transport Process; Validation; analytical method; cell motility; clinical application; controlled release; design; granulocyte; in vivo; in vivo Model; inhibitor/antagonist; innovation; killings; microbial; microorganism; migration; neutrophil; novel; pathogen; pre-clinical; prevent; public health relevance; response; secretory protein; small molecule; trafficking; treatment strategy

DESCRIPTION (provided by applicant): Intracellular vesicular transport is essential for nearly all aspects of neutrophil physiology and defects in this mechanism leads to disease in humans. In neutrophils, vesicular trafficking is directly associated with the processes of exocytosis, phagocytosis, adhesion and chemotaxis; however, the molecular mechanisms that regulate mobilization of the different neutrophil secretory organelles during secretion, pathogen internalization or migration are incompletely understood. We have identified several key regulatory factors essential for the control of granule trafficking in neutrophils including the smll GTPase Rab27a that controls the mobilization and secretion of neutrophil granules in vitro and in vivo. This function is regulated by two Rab27a effectors, JFC1 and Munc13-4. Furthermore, we have recently identified previously unrecognized neutrophil secretory factors that interact with JFC1 and regulate cytoskeleton remodeling, vesicular trafficking and exocytosis and have pioneered the development of systems biology approaches to analyze vesicular dynamics and actin remodeling in granulocytes. In this application, we propose to use innovative analytical and quantitative methods to elucidate the mechanisms regulating vesicular transport associated with exocytosis, phagocytosis and chemotaxis in neutrophils. We also propose to use novel small molecule inhibitors of Rab27a-effector binding to interrogate mechanisms of vesicular transport and elucidate Rab27a-dependent processes in disease using an in vivo model of endotoxin-induced systemic inflammation. Since the toxic content of neutrophil granules is injurious to the host and neutrophil secretory proteins play a fundamental role in the damage to endothelium associated with endotoxemia and sepsis, these studies have important physiological significance and potential clinical applications in disease. We hypothesize that the differential regulation of vesicular transport by Rab27a and its effectors is an essential mechanism to determine specific neutrophil functions and responses to insult. We also propose that small molecule modulators of vesicular transport pathways will prevent some of the deleterious consequences of neutrophil secretion during systemic inflammation. To test our hypotheses we propose the following Specific Aims: 1) To define the mechanisms that regulate vesicular trafficking and actin remodeling during exocytosis in granulocytes and establish the role of the Rab27a effector JFC1 in this mechanism; 2) To establish the mechanisms regulated by Munc13-4 in vesicular transport and vesicular fusion associated with exocytosis, phagocytosis and bacterial killing; 3) To test the hypothesis that novel small molecule inhibitors of Rab27a-effector binding modulate neutrophil granule trafficking and exocytosis and attenuate inflammation. The results of the proposed research plan should uncover the molecular mechanisms regulating vesicular transport in neutrophils and lead to effective molecular strategies for the treatment of inflammatory diseases.

描述（由申请人提供）：细胞内囊泡转运对中性粒细胞生理学的几乎所有方面都是必不可少的，该机制的缺陷导致人类疾病。在中性粒细胞中，囊泡运输与胞吐、吞噬、粘附和趋化过程直接相关;然而，在分泌、病原体内化或迁移过程中调节不同中性粒细胞分泌细胞器动员的分子机制尚不完全清楚。我们已经确定了几个关键的调节因子的控制颗粒运输的嗜中性粒细胞，包括smll GTCRab 27 a，控制动员和分泌的嗜中性粒细胞颗粒在体外和体内。该功能由两个Rab27a效应子JFC 1和Munc13 - 4调节。此外，我们最近已经确定了以前未被识别的中性粒细胞分泌因子与JFC 1相互作用，调节细胞骨架重塑，囊泡运输和胞吐作用，并率先发展系统生物学方法来分析囊泡动力学和肌动蛋白重塑粒细胞。在这个应用中，我们建议使用创新的分析和定量方法来阐明调节与中性粒细胞的胞吐，吞噬和趋化性相关的囊泡运输的机制。我们还建议使用Rab27a效应器结合的新型小分子抑制剂来询问囊泡转运机制，并使用内毒素诱导的全身性炎症的体内模型来阐明Rab27a依赖性疾病过程。由于中性粒细胞颗粒的毒性成分对宿主具有损伤作用，中性粒细胞分泌蛋白在内毒素血症和脓毒症相关的内皮损伤中起重要作用，因此这些研究具有重要的生理意义和潜在的临床应用价值。我们推测Rab27a及其效应子对囊泡转运的差异调节是确定特定中性粒细胞功能和对损伤反应的重要机制。我们还提出，囊泡转运途径的小分子调节剂将防止一些系统性炎症过程中中性粒细胞分泌的有害后果。为了验证我们的假设，我们提出了以下具体目标：1）确定在粒细胞胞吐过程中调节囊泡运输和肌动蛋白重塑的机制，并确定Rab27 a效应子JFC 1在该机制中的作用; 2）确定Munc13 - 4在囊泡运输和囊泡融合中与胞吐、吞噬和细菌杀伤相关的调节机制; 3）检验Rab27 a效应物结合的新型小分子抑制剂调节嗜中性粒细胞颗粒运输和胞吐并减轻炎症的假设。拟议的研究计划的结果应该揭示调节中性粒细胞囊泡转运的分子机制，并导致有效的治疗炎症性疾病的分子策略。