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

• 批准号: 10582013
• 负责人: Scott David Hansen
• 金额: $ 7.86万
• 依托单位: UNIVERSITY OF OREGON
• 依托单位国家: 美国
• 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/10582013
• 关键词: Bacteria; Behavior; Biochemistry; Biophysics; Biosensor; Cell Surface Receptors; Cell membrane; Cell physiology; Cells; Cellular biology; Chemicals; Clustered Regularly Interspaced Short Palindromic Repeats; Coin; Communication; Cues; Decision Making; Environment; Enzymes; Goals; Guanosine Triphosphate Phosphohydrolases; Human; Immune; Immunomodulators; In Vitro; Infection; Inflammation; Innate Immune System; Knowledge; Leukocytes; Lipids; Malignant Neoplasms; Membrane; Molecular; Monomeric GTP-Binding Proteins; Movement; Pathogenicity; Phosphatidylinositol Phosphates; Phosphoric Monoester Hydrolases; Proteins; Regulation; Research Personnel; Role; Signal Transduction; Signaling Molecule; Source; Techniques; Technology; Travel; Virus; base; cancer cell; cell motility; combat; fluorescence imaging; genome editing; live cell imaging; molecular imaging; neutrophil; novel therapeutics; optogenetics; reconstitution; recruit; scaffold; single molecule; tool

Abstract 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.

抽象的 免疫细胞解释其环境中的化学信号并做出控制其免疫细胞的决定 命运。例如，人类中性粒细胞通常通过极化和迁移来响应信号 朝向化学源。这些细胞功能的关键是之间的动态相互作用 细胞表面受体、小 GTP 酶和合成磷脂酰肌醇的酶 磷酸盐（PIP）脂质。本研究旨在破译控制通信的机制 质膜上这些不同类别的信号分子之间的关系。利用人类 中性粒细胞中，我们发现 Cdc42 GTPase 和称为 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。 这种行为被称为兴奋性，涉及蛋白质募集开和关的重复循环 膜。本研究的目的是确定 SHIP1 如何调节兴奋性 通过整合来自脂质和膜束缚蛋白的信号来构建信号网络。 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 膜缔合和磷酸酶活性（目标 1）。利用调节因素 细胞中的兴奋信号网络，我们将重建控制SHIP1的机制 膜募集、自抑制释放和激活（目标 2）。同时，我们将使用 基于 CRISPR 的基因组编辑、光遗传学和荧光定量活细胞成像 生物传感器阐明 PIP 脂质和小 GTP 酶之间的通讯如何受到调节 由 SHIP1 提供。使用这些工具，我们将确定 SHIP1 作为信令网络的角色 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 和细胞生物学来解释 PIP 脂质、小 GTPases 和 SHIP1 如何协同控制 中性粒细胞的兴奋信号网络和细胞迁移。通过揭开白血如何 细胞感知、解释和响应致病信号我们将填补知识空白 关于这些信号分子如何协调细胞运动与潜在的 兴奋网络。这一发现可能为研究人员开发新疗法打开大门 可用于调节免疫细胞功能，对抗感染、炎症、 和癌症。