Mechanisms mediating immune response upon sensing of nuclear viral DNA

感测核病毒 DNA 介导免疫反应的机制

• 批准号: 9027921
• 负责人: Ileana M. Cristea
• 金额: $ 31.87万
• 依托单位: PRINCETON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-09-15 至 2019-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9027921
• 关键词: Acute Promyelocytic Leukemia; Address; Antiviral Agents; Antiviral Response; Antiviral Therapy; Belief; Binding; Biochemistry; Biological Assay; Cell Nucleus; Cells; Cytomegalovirus; Cytomegalovirus Infections; Cytoplasm; DNA; DNA Viruses; Development; Drug Design; Drug Targeting; Endoplasmic Reticulum; Equilibrium; Event; Fluorescent in Situ Hybridization; Hand; Health; Herpesviridae; Herpesvirus 1; Homo; Human; Hybrids; Immune; Immune System Diseases; Immune response; Immune system; Immunity; Infection; Interferons; Lead; Life; Light; Maintenance; Malignant Neoplasms; Mammalian Cell; Mediating; Methods; Microbe; Microscopy; Molecular Biology; Morbidity - disease rate; Nuclear; Nuclear Export; Pathway interactions; Pilot Projects; Property; Proteins; Proteomics; RNA Viruses; Recruitment Activity; Regulation; Replication Origin; Research; Role; Series; Signal Transduction; Staging; System; Testing; Ubiquitin; Viral Proteins; Virus; Work; adapter protein; base; cellular imaging; cytokine; interdisciplinary approach; marenostrin; mortality; novel; novel therapeutics; optogenetics; pathogen; public health relevance; sensor; small hairpin RNA; transmission process; viral DNA

 DESCRIPTION (provided by applicant): The ability of the mammalian immune system to recognize pathogenic DNA, such as DNA from viruses, is essential for the onset of intrinsic and innate immune responses. This recognition is accomplished by specialized cellular proteins called DNA sensors, which bind foreign DNA and elicit the secretion of cytokines to alert neighboring cells and inhibit the spread of infection. Until recently, the sensing of foreign DNA was thought to occur only in subcellular compartments normally devoid of DNA. However, this long-standing belief failed to explain how the cell detects nuclear-replicating DNA viruses. Our lab's recent characterization of the first identified nuclear DNA sensor, IFI16, has helped to firmly establish the concept of nuclear sensing. We demonstrated that IFI16 functions to sense herpesviruses, including the important pathogens, human cytomegalovirus (HCMV) and herpes simplex virus type 1 (HSV-1) (Li et al. PNAS 2012; Li et al. Cell Host Microbe 2013). The discovery that sensing can occur in the nucleus opens a new direction for research in immunity that will increase our understanding of how balanced immune responses work to maintain a healthy system and how their misregulation leads to immune disorders, cancers, and virus-induced morbidity and mortality. However, a fundamental question that has yet to be answered is how the immune signal is propagated from the nucleus. We found that, while IFI16 remains nuclear during the early stages of HCMV and HSV-1 infections, the IFI16-mediated induction of antiviral cytokines requires the endoplasmic reticulum adapter protein STING-a hub for DNA sensing pathways. The requirements for eliciting an immune response from the nucleus and the mechanism of immune signal propagation to the cytoplasm remain elusive. Our proposal will address these important questions. First, we will characterize the mechanism by which IFI16 rapidly co-localizes with viral DNA in the nucleus. We have discovered that IFI16 is recruited to promyelocytic leukemia (PML) nuclear bodies (NBs) following infection. PML-NBs have been implicated in antiviral response and shown to localize to origins of replication of herpesviruses. Using a multidisciplinary approach, we will test our hypothesis that PML-NBs function as interaction centers where IFI16 concentrates its binding of viral DNA, setting the stage for immune signal initiation. Second, we will use molecular biology, biochemistry, optogenetics, and live cell imaging to define the IFI16 properties required for initiating nuclear immune signals. We hypothesize that signal initiation requires IFI16 oligomerization via its pyrin domain at PML-NBs. Third, we will determine how the nucleus-derived immune signal is transmitted to the cytoplasm. We discovered that IFI16 interacts with interferon-inducible IFIT proteins, which shuttle to the nucleus upon infection. We propose mechanistic studies to test our hypothesis that IFI16-dependent immune signaling is relayed by such interactions to the STING hub. Collectively, our results will characterize a newly discovered aspect of immunity, nuclear sensing, by defining the fundamental mechanisms required for immune signal propagation.

 描述（由申请人提供）：哺乳动物免疫系统识别病原性DNA（例如来自病毒的DNA）的能力对于内在和先天免疫反应的发生至关重要。这种识别是通过称为 DNA 传感器的特殊细胞蛋白来完成的，它结合外源 DNA 并引发细胞因子的分泌，以警告邻近细胞并抑制感染的传播。直到最近，外源 DNA 的感知还被认为只发生在通常没有 DNA 的亚细胞区室中。然而，这种长期存在的信念未能解释细胞如何检测核复制 DNA 病毒。我们实验室最近对第一个确定的核 DNA 传感器 IFI16 进行了表征，有助于牢固地确立核传感的概念。我们证明了 IFI16 具有感知疱疹病毒的功能，包括重要的病原体、人巨细胞病毒 (HCMV) 和 1 型单纯疱疹病毒 (HSV-1)（Li 等人，PNAS 2012；Li 等人，Cell Host Microbe，2013）。细胞核中发生传感的发现为免疫研究开辟了一个新方向，这将增加我们对平衡免疫反应如何维持健康系统以及免疫反应失调如何导致免疫紊乱、癌症以及病毒引起的发病和死亡的理解。然而，一个尚未解答的基本问题是免疫信号如何从细胞核传播。我们发现，虽然 IFI16 在 HCMV 和 HSV-1 感染的早期阶段仍保留在细胞核中，但 IFI16 介导的抗病毒细胞因子的诱导需要内质网接头蛋白 STING——DNA 传感途径的枢纽。从细胞核引发免疫反应的要求以及免疫信号传播到细胞质的机制仍然难以捉摸。我们的提案将解决这些重要问题。首先，我们将描述 IFI16 与病毒 DNA 在细胞核中快速共定位的机制。我们发现 IFI16 在感染后被招募到早幼粒细胞白血病 (PML) 核体 (NB) 中。 PML-NB 与抗病毒反应有关，并显示出定位于疱疹病毒的复制起点。我们将使用多学科方法来检验我们的假设，即 PML-NB 充当相互作用中心，其中 IFI16 集中结合病毒 DNA，为免疫信号启动奠定基础。其次，我们将利用分子生物学、生物化学、光遗传学和活细胞成像来定义启动核免疫信号所需的 IFI16 特性。我们 假设信号启动需要 IFI16 通过 PML-NB 上的 Pyrin 结构域进行寡聚化。第三，我们将确定核源性免疫信号如何传递到细胞质。我们发现 IFI16 与干扰素诱导的 IFIT 蛋白相互作用，后者在感染后穿梭至细胞核。我们提出机制研究来检验我们的假设，即 IFI16 依赖性免疫信号通过这种相互作用传递到 STING 中枢。总的来说，我们的结果将通过定义免疫信号传播所需的基本机制来表征免疫的新发现的方面，即核传感。