The adaptor protein Crk in immune responses

免疫反应中的接头蛋白 Crk

基本信息

批准号：
9428868
负责人：
Dongfang Liu
金额：
$ 53.15万
依托单位：
METHODIST HOSPITAL RESEARCH INSTITUTE
依托单位国家：
美国
项目类别：
财政年份：
2017
资助国家：
美国
起止时间：
2017-03-09 至 2018-02-12
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9428868
关键词：
Actins Adaptor Signaling Protein Affect Algorithms Apoptosis Autologous Automobile Driving Biochemical Biological Models Biological Response Modifiers Birth CRKL protein Cell Line Cell physiology Cell-Mediated Cytolysis Cells Chronic Disease Clinical Complex Custom Cytoplasmic Granules Cytoskeleton Data DiGeorge Syndrome Disease Effector Cell Energy Transfer Event Explosion Fluorescence Glioblastoma Goals Hematologic Neoplasms Host Defense Human Image Imaging Techniques Imaging technology Immune Immune response Immune system Immunity Immunologic Deficiency Syndromes Immunologics Immunology Immunoprecipitation In Vitro Infection Killer Cells Knockout Mice Knowledge Lipid Bilayers Lytic Malignant Neoplasms Malignant neoplasm of lung Malignant neoplasm of ovary Microscopy Modeling Molecular Molecular Conformation NK Cell Activation Natural Killer Cells Patients Phosphorylation Phosphotransferases Play Protein Family Proteins Recruitment Activity Regulation Research Resolution Role Shapes Signal Transduction Signaling Molecule Small Interfering RNA Stem cells Structure Synapses System Techniques Testing Tissues Transfection Viral Cancer Virus Virus Diseases Work biological systems cell killing cell mediated immune response congenital immunodeficiency cytotoxic experimental study fighting gene therapy immunological synapse immunoregulation in vivo insight kinase inhibitor knock-down live cell microscopy malignant breast neoplasm molecular imaging novel novel therapeutics population based protein function proto-oncogene protein c-crk receptor sarcoma single molecule spatiotemporal

项目摘要

Project Summary Natural killer (NK) cells play an important role in the human immune response to infection and malignancy. How these cells effectively distinguish between diseased and healthy tissue is one of the key unsolved problems in immunology today. The proposed work seeks to identify the mechanism(s) by which the small adaptor protein CT10 regulator of kinase (Crk), and its phosphorylation, control NK cell activation and inhibition by using both human NK cells and novel NK cell-specific conditional knockout mice. The long-term goal is to use this knowledge and the novel imaging techniques developed herein to uncover the molecular basis of NK cell activation and inhibition, and to develop new treatments for human primary immunodeficiency diseases and chronic diseases such as cancer and viral infection. NK cells kill target cells through the polarized release of lytic granules through a specialized region of cell-cell contact known as the immunological synapse (IS). Through previous studies of the cytotoxic (Liu, D. et al., Immunity, 2009, Cover Article) and inhibitory (Liu, D. et al., Immunity, 2012) IS, we discovered that Crk plays an essential upstream role at the IS, influencing signaling events required for both activation and inhibition. The molecular mechanisms underlying this dual role, however, remain unclear. We hypothesize that receptor-driven phosphorylation of Crk acts as a molecular switch, driving a conformational change, which in turn determines Crk's ability to interact with critical downstream signaling molecules and ultimately shapes the actin cytoskeleton into a functional IS. Guided by strong preliminary data, we will test these hypotheses via three Specific Aims: 1) Define the precise molecular mechanisms by which Crk controls NK cell activation and inhibition. The proposed work capitalizes on the recent explosion in single molecule imaging and will bring cutting-edge single molecule imaging technology to the field of NK research, with a focus on human primary immunodeficiency diseases; 2) Determine the role of Crk-like protein (CrkL) in NK cells from patients with partial DiGeorge syndrome (pDGS). Using one of most common (1 in 4,000 births) immunodeficiency diseases, pDGS (mainly caused by CrkL haploinsufficiency), we will determine how loss of CrkL function affects NK cell-mediated cytotoxicity in pDGS and its molecular mechanisms; 3) Determine whether Crk is required for NK function in vivo. Leveraging novel NK cell-specific Crk knockout mice that we have already generated, we will determine the roles that Crk molecules play in NK cell-mediated immune responses to viral infection and cancer in vivo. The proposed work involves key signaling players and novel regulatory mechanisms and is broadly relevant with direct clinical implications for the treatment of primary immunodeficiency diseases and cancer because elevated expression and increased phosphorylation of Crk are common to various cancers (e.g., lung cancer, breast cancer, glioblastoma, sarcomas, ovarian cancer, and hematopoietic cancers). The proposed work also generates a novel model system in which to determine Crk's role as a master regulatory molecule. !

项目概要自然杀伤 (NK) 细胞在人体对感染和恶性肿瘤的免疫反应中发挥着重要作用。这些细胞如何有效区分患病组织和健康组织是未解决的关键问题之一当今免疫学的问题。拟议的工作旨在确定小型企业的机制接头蛋白 CT10 激酶 (Crk) 调节因子，及其磷酸化，控制 NK 细胞的激活和抑制通过使用人类 NK 细胞和新型 NK 细胞特异性条件敲除小鼠。长期目标是利用这些知识和本文开发的新型成像技术来揭示 NK 的分子基础细胞激活和抑制，并开发人类原发性免疫缺陷疾病的新疗法以及癌症和病毒感染等慢性疾病。 NK细胞通过极化释放杀死靶细胞通过称为免疫突触 (IS) 的细胞与细胞接触的特殊区域来释放裂解颗粒。通过之前对细胞毒性（Liu, D. et al., Immunity, 2009, Cover Article）和抑制性（Liu, D. et al., Immunity, 2012) IS，我们发现 Crk 在 IS 中起着重要的上游作用，影响激活和抑制所需的信号事件。这种双重作用背后的分子机制然而，其作用仍不清楚。我们假设受体驱动的 Crk 磷酸化充当分子开关，驱动构象变化，进而决定 Crk 与关键蛋白相互作用的能力下游信号分子并最终将肌动蛋白细胞骨架塑造成功能性 IS。指导者有了强有力的初步数据，我们将通过三个具体目标来测试这些假设：1）定义精确的 Crk 控制 NK 细胞激活和抑制的分子机制。拟议的工作利用最近单分子成像的爆炸性发展，将带来尖端的单分子将成像技术引入NK研究领域，重点关注人类原发性免疫缺陷疾病； 2）确定部分 DiGeorge 综合征患者 NK 细胞中 Crk 样蛋白 (CrkL) 的作用（pDGS）。使用最常见的免疫缺陷疾病之一（每 4,000 名新生儿中就有 1 名）pDGS（主要由 CrkL 单倍体不足），我们将确定 CrkL 功能的丧失如何影响 NK 细胞介导的细胞毒性 pDGS及其分子机制； 3) 确定体内NK功能是否需要Crk。利用我们已经生成的新型 NK 细胞特异性 Crk 敲除小鼠，我们将确定 Crk 分子在 NK 细胞介导的体内病毒感染和癌症免疫反应中发挥的作用。这拟议的工作涉及关键信号参与者和新颖的监管机制，并且与广泛相关对原发性免疫缺陷疾病和癌症的治疗具有直接的临床意义，因为 Crk 表达升高和磷酸化增加对于多种癌症（例如肺癌、乳腺癌、胶质母细胞瘤、肉瘤、卵巢癌和造血系统癌症）。拟议的工作还生成了一个新颖的模型系统，以确定 Crk 作为主调节分子的作用。！