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Deciphering the mechanism of SHIP1 regulation in human neutrophils

破译 SHIP1 在人类中性粒细胞中的调节机制

基本信息

批准号：
10623312
负责人：
Scott David Hansen
金额：
$ 30.14万
依托单位：
UNIVERSITY OF OREGON
依托单位国家：
美国
项目类别：
财政年份：
2021
资助国家：
美国
起止时间：
2021-07-01 至 2026-05-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10623312
关键词：
Actins Architecture Bacteria Behavior Binding Biochemical Biochemistry Biological Models Biophysics Biosensor C-terminal CRISPR interference Cell Line Cell Polarity Cell Surface Receptors Cell membrane Cell physiology Cells Cellular biology Chemicals Chemotaxis Clustered Regularly Interspaced Short Palindromic Repeats Coin Communication Cues Decision Making Defect Engineering Environment Enzymes Exhibits Fluorescence Microscopy Genetic Goals Guanosine Triphosphate Phosphohydrolases Human Immune Immune response Immune system Immunologic Receptors In Vitro Infection Inflammation Innate Immune System Invaded Knowledge Leukocytes Lipid Bilayers Lipids Malignant Neoplasms Membrane Molecular Monomeric GTP-Binding Proteins Movement PIK3CG gene Pathogenicity Phenotype Phosphatidylinositol Phosphates Phosphatidylinositols Phosphatidylserines Phosphoric Monoester Hydrolases Proteins Receptor Activation Regulation Reporting Research Research Personnel Role Scaffolding Protein Signal Transduction Signaling Molecule Source System Techniques Technology Testing Travel Virus Visualization cancer cell cell motility chemical release combat fMet-Leu-Phe receptor fluorescence imaging genome editing immune modulating agents immunoregulation inhibitor inorganic phosphate live cell imaging migration molecular imaging mutant neutrophil novel therapeutics optogenetics pathogen phosphoinositide-3,4,5-triphosphate phosphoinositide-3,4-bisphosphate reconstitution recruit scaffold single molecule tool

项目摘要

Research Summary Statement Immune cells interpret chemical cues in their environment and make decisions that control their fate. For example, human neutrophils often respond to signals by polarizing and migrating toward the chemical source. Critical for these cellular functions is the dynamic interplay between cell surface receptors, small GTPases, and the enzymes that synthesize phosphatidylinositol phosphate (PIP) lipids. This study aims to decipher the mechanisms the control communication between these different classes of signaling molecules at the plasma membrane. Using human neutrophils, we find that Cdc42 GTPase and a lipid phosphatase denoted SHIP1 trigger a molecular signal that propagates across the plasma membrane in the form of a traveling wave. This behavior, coined excitability, involves repetitive cycles of protein recruitment ON and OFF the membrane. The goal of this study is to determine how SHIP1 regulates the excitable signaling network by integrating signals derived from lipids and membrane tethered proteins. Using a variety of in vitro biochemistry techniques, including supported membrane technology and single molecule imaging, we will determine how lipid composition controls SHIP1 membrane association and phosphatase activity (Aim 1). Using factors that regulate the excitable signaling network in cells, we will reconstitute mechanisms that control SHIP1 membrane recruitment, release of autoinhibition, and activation (Aim 2). In parallel, we will use CRISPR based genome editing, optogenetics, and quantitative live cell imaging of fluorescent biosensors to elucidate how communication between PIP lipids and small GTPase is regulated by SHIP1. Using these tools we will determine the role SHIP1 serves as signaling network scaffold versus a lipid phosphatase (Aim 3). Overall, this study will unify membrane biophysics and cell biology to explain how PIP lipids, small GTPases, and SHIP1 synergistically control the excitable signaling network and cell migration in neutrophils. By unraveling how white blood cells sense, interpret, and respond to pathogenic signals we will fill a gap in knowledge concerning how these signaling molecules coordinate cellular movement with the underlying excitable network. This discovery could open doors for researchers to develop new therapeutics that can be used to modulate immune cell functions in ways that combat infection, inflammation, and cancer.

研究总结声明免疫细胞解释环境中的化学信号并做出控制其命运的决定。为了例如，人类中性粒细胞通常通过极化并向化学源迁移来响应信号。这些细胞功能的关键是细胞表面受体、小 GTP 酶和合成磷脂酰肌醇磷酸 (PIP) 脂质的酶。本研究旨在破译等离子体中这些不同类别的信号分子之间的控制通信机制膜。使用人类中性粒细胞，我们发现 Cdc42 GTPase 和表示为 SHIP1 的脂质磷酸酶触发以行波形式穿过质膜传播的分子信号。这行为，即兴奋性，涉及膜上和膜外蛋白质募集的重复循环。这本研究的目标是确定 SHIP1 如何通过整合信号来调节可兴奋信号网络源自脂质和膜束缚蛋白。使用各种体外生物化学技术，包括支持膜技术和单分子成像，我们将确定脂质成分如何控制 SHIP1 膜关联和磷酸酶活性（目标 1）。利用调节兴奋的因素细胞内的信号网络，我们将重建控制SHIP1膜招募、释放的机制自抑制和激活（目标 2）。与此同时，我们将使用基于 CRISPR 的基因组编辑、光遗传学、荧光生物传感器的定量活细胞成像，以阐明 PIP 脂质之间的通讯方式小GTP酶受SHIP1调节。使用这些工具，我们将确定 SHIP1 作为信号传递的角色网络支架与脂质磷酸酶（目标 3）。总的来说，这项研究将统一膜生物物理学和细胞生物学解释 PIP 脂质、小 GTPases 和 SHIP1 如何协同控制兴奋信号传导中性粒细胞的网络和细胞迁移。通过揭示白细胞如何感知、解释和响应致病信号，我们将填补关于这些信号分子如何协调细胞的知识空白与底层可兴奋网络的运动。这一发现可能为研究人员开发新产品打开大门可用于调节免疫细胞功能以对抗感染的新疗法，炎症、癌症。