The mechanisms underlying the GPCR-mediated chemotaxis in D. discoideum

D. discoideum GPCR 介导的趋化机制

• 批准号: 9566620
• 负责人: Tian Jin
• 金额: $ 61.81万
• 依托单位: NATIONAL INSTITUTE OF ALLERGY AND INFECTIOUS DISEASES
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/9566620
• 关键词: Actins; Amoeba genus; Animal Model; Bacteria; Bacterial Infections; Binding; Biochemical; Biological Models; Blood Circulation; Cells; Chemicals; Chemotactic Factors; Chemotaxis; Computer Assisted; Coupled; Cyclic AMP; Cyclic AMP Receptors; Cytoskeleton; Cytosol; Data Set; Defect; Dictyostelium discoideum; Dissociation; Escherichia coli; Eukaryotic Cell; F-Actin; Folic Acid; Food; G-Protein-Coupled Receptors; G-substrate; GTP-Binding Protein alpha Subunits, Gs; GTP-Binding Proteins; Glutaminase; Goals; Heterotrimeric GTP-Binding Proteins; Hormones; Human; Human body; Immune; Immunology; Impairment; Infection; Investigation; Knock-out; Light; Mediating; Membrane; Modeling; Molecular; Movement; Natural Immunity; Neurotransmitters; Organism; Phagocytes; Phagocytosis; Physiological; Process; Proteins; Recruitment Activity; Signal Pathway; Signal Transduction; Signal Transduction Pathway; Site; Source; Stimulus; Surface; Techniques; base; cell motility; chemokine; directional cell; extracellular; fMet-Leu-Phe receptor; fighting; fluorescence imaging; genome database; imaging modality; immune phagocytosis; macrophage; migration; neutrophil; novel; particle; pathogen; phosphoproteomics; polymerization; receptor-mediated signaling; response; spatiotemporal

1: Eukaryotic cells chemotax in a wide range of chemoattractant concentration gradients, and thus need inhibitory processes that terminate cell responses to reach adaptation while maintaining sensitivity to higher-concentration stimuli. However, the molecular mechanisms underlying inhibitory processes are still poorly understood. Here, we reveal a locally controlled inhibitory process in a GPCR-mediated signaling network for chemotaxis in Dictyostelium discoideum. We discover a novel negative regulator of Ras signaling, C2GAP1, which localizes at the leading edge of chemotaxing cells, is activated by and is essential for GPCR-mediated Ras adaptation. We show that both C2 and GAP domains are required for the membrane targeting of C2GAP1, and that GPCR-triggered Ras activation recruits C2GAP1 from cytosol and retains it on the membrane to locally inhibit Ras signaling. The altered Ras activation results in impaired gradient sensing and excessive polymerization of F-actin in c2gap1 knockout (c2gap1-) cells, leading to chemotaxis defects. Remarkably, c2gap1- cells display altered cell response, impaired directional sensing, and chemotaxis defects in a chemoattractant concentration-dependent fashion. Thus, we have uncovered a novel inhibitory mechanism required for the adaptation and long-range chemotax.(Xu et al., in revision). 2: Human phagocytes, including neutrophils and macrophage, are an essential part of innate immunity. Upon bacterial infection, neutrophils leaves circulation and migration to infection sites to fight pathogens. They seek bacteria by detecting chemoattractants generated from the bacterial infection site and chase them via chemotaxis. Once reaching the site, they bind and ingest bacteria by recognizing signals form the bacterial surface via phagocytosis. Dictyostelium discoideum amoeba are professional phagocytes that track down bacteria by chemotaxis and capture and ingest them as food through phagocytosis. Studies in the simple model organism of D. discoideum have made tremendous contribution to our current understanding of molecular mechanisms underlying chemotaxis and phagocytosis of phagocytes. Recently, we discovered that D. discoideum amoeba use a chemoattractant GPCR fAR1 to detect folic acid released from bacteria for both chemotaxis to catch bacteria and phagocytosis to ingest them (Pan et al, 2016). This finding suggest to us that a chemoattractant GPCR-mediated signaling network controls reorganization of the actin cytoskeleton for both chemotaxis and phagocytosis, which represents a paradigm-shifting new concept in immunology. Thus, investigation of molecular components involved in chemotaxis and (or) phagocytosis in D. discoideum will continually shed light on the molecular mechanisms controlling migration of immune cells and as well as phagocytosis by immune cells to eliminate bacterial pathogens from human body. Recently, we discovered that formyl-peptide receptors (fPR GPCR) coupled with heterotrimeric Gi proteins mediate chemotaxis as well as phagocytosis of fMLP-coated particles and E. coli. Our studies revealed an evolutionarily conserved mechanism that directs professional phagocytes migrating toward bacteria via chemotaxis and promotes them to engulf bacterial via surface phagocytosis as an essential part of innate immunity.

一曰：真核细胞在广泛的化学引诱物浓度梯度中趋化，因此需要终止细胞反应以达到适应的抑制过程，同时保持对较高浓度刺激的敏感性。然而，抑制过程的分子机制仍然知之甚少。在这里，我们揭示了一个局部控制的抑制过程中GPCR介导的信号网络的趋化性Dictyosteelium discoideum。我们发现了一种新的Ras信号负调控因子C2 GAP 1，它位于趋化细胞的前沿，被GPCR介导的Ras适应激活，并且是必需的。我们发现，C2和GAP结构域都是C2 GAP 1的膜靶向所必需的，并且GPCR触发的Ras激活从胞质溶胶中招募C2 GAP 1并将其保留在膜上以局部抑制Ras信号传导。Ras激活的改变导致c2 gap 1敲除细胞中梯度感应受损和F-肌动蛋白过度聚合，导致趋化性缺陷。值得注意的是，c2 gap 1-细胞显示改变的细胞反应，受损的方向感，和趋化缺陷的化学引诱物浓度依赖性的方式。因此，我们已经发现了一种新的抑制机制所需的适应和远程chemotax。(Xu例如，修订中）。 2：人类吞噬细胞，包括中性粒细胞和巨噬细胞，是先天免疫的重要组成部分。 在细菌感染后，中性粒细胞离开循环并迁移到感染部位以对抗病原体。 它们通过检测从细菌感染部位产生的化学引诱物来寻找细菌，并通过趋化性来追逐它们。 一旦到达该位点，它们通过吞噬作用识别来自细菌表面的信号来结合和摄取细菌。 盘状网囊藻变形虫是一种专职吞噬细胞，通过趋化作用追踪细菌，并通过吞噬作用捕获和摄取细菌作为食物。 本文对D. discoideum对我们目前对吞噬细胞趋化性和吞噬作用的分子机制的理解做出了巨大贡献。 最近，我们发现D. discoideum amoeba使用化学引诱剂GPCR fAR 1来检测细菌释放的叶酸，以实现捕获细菌的趋化作用和吞噬细菌的吞噬作用（Pan等人，2016）。 这一发现向我们表明，趋化GPCR介导的信号网络控制肌动蛋白细胞骨架的重组，用于趋化和吞噬作用，这代表了免疫学中的一个范式转变的新概念。 因此，对D. discoideum将继续揭示控制免疫细胞迁移的分子机制，以及免疫细胞的吞噬作用，以消除人体内的细菌病原体。 最近，我们发现与异源三聚体Gi蛋白偶联的甲酰肽受体（fPR GPCR）介导fMLP包被颗粒和E.杆菌 我们的研究揭示了一个进化上保守的机制，指导专业吞噬细胞通过趋化性向细菌迁移，并促进它们通过表面吞噬作用吞噬细菌，作为先天免疫的重要组成部分。