Project 2: Genes required for dendritic cell responses to pathogens and T cells

项目2：树突状细胞对病原体和T细胞做出反应所需的基因

• 批准号: 10207350
• 负责人: Nir Hacohen
• 金额: $ 69.08万
• 依托单位: HARVARD MEDICAL SCHOOL
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-07-05 至 2023-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10207350
• 关键词: Acute; Address; Animals; Antigen Presentation; Antigens; Attention; Bacteria; Binding; Biological Assay; Bone Marrow; CD14 Antigen; CD14 gene; CD4 Positive T Lymphocytes; CRISPR library; CRISPR screen; CRISPR/Cas technology; Candidate Disease Gene; Caspase; Cell Communication; Cell physiology; Cells; Cellular biology; Chromatin; Clustered Regularly Interspaced Short Palindromic Repeats; Collaborations; Communication; Communities; Cross Presentation; Dendritic Cells; Dendritic cell activation; Detection; Endocytosis; Endosomes; Event; Genes; Genetic; Genetic Screening; Genetic Transcription; Genome; Grant; Helper-Inducer T-Lymphocyte; Histones; Host Defense; Human; Immune; Immune system; Immunity; Infection; Inflammation; Inflammatory; Informatics; Ingestion; Innate Immune Response; Innate Immune System; Interleukin-1; Interleukin-2; Investigation; Knowledge; Lead; Lipids; Lipopolysaccharides; Lymphocytic choriomeningitis virus; Lymphoid; Mammalian Cell; Measurement; Membrane Proteins; Messenger RNA; Microbe; Monitor; Mus; Pathway interactions; Phosphorylation; Physiology; Play; Process; Protein Biosynthesis; Proteins; Proteome; RNA Splicing; RNA chemical synthesis; Regulation; Reporting; Research; Research Personnel; Resolution; Role; Series; Signal Pathway; Signal Transduction; Signaling Protein; Site; Sorting - Cell Movement; System; T-Cell Activation; T-Lymphocyte; TCF Transcription Factor; TLR4 gene; Transgenic Mice; Ubiquitination; Work; acute infection; adaptive immunity; base; candidate selection; cell type; cytokine; gene discovery; gene function; genetic regulatory protein; genome editing; genome-wide; in vivo; innovation; knockout gene; macrophage; microbial; novel; pathogen; receptor; response; single-cell RNA sequencing; transcription factor

The dendritic cells (DCs) of the mammalian innate immune system are responsible for detection of microbial encounters and the initiation of inflammation and adaptive immunity. This central role of DCs in host physiology has attracted much attention, and these cells are the subject of active investigation. Despite the appreciation of the importance of DCs, there remain large and fundamental gaps in our knowledge of their mechanisms of regulation. For instance, DCs encode several receptors that detect bacterial lipopolysaccharides (LPS), yet the functions of only one (TLR4) have been examined extensively. Recent work by us and others have identified the LPS receptors CD14 and caspase-11 as being capable of inducing novel signaling pathways that proceed either upstream of TLR4 (CD14) or in parallel to TLR4 (caspase-11). The collective actions of CD14, TLR4 and caspase-11 are important for DC activation and host defense, yet their mechanisms of action are poorly defined. Thus, significant gaps in our knowledge exist to explain the earliest stages of DC interactions with bacteria. We also lack an understanding of how DCs interact with T cells at later stages of infection, yet it is clear that these interactions lead to profound changes in the activities of both cell types. Much of the research into these interactions has focused on how DCs promote changes in T cell activities. In contrast, we have a minimal understanding of how T cells influence the activity of DCs. In this application, we propose to fill these gaps in our knowledge of DC biology through forward genetic screening for novel regulatory factors in mice. This approach is facilitated by recent advances in genome editing provided by CRISPR-based technologies. A pipeline of gene discovery will be generated through the use of established and emerging FACS-based assays. These assays will be used in vivo and ex vivo to identify DCs that are deficient for regulators of DC interactions with microbes or T cells. All screens will depend on significant interactions between the informatics, mouse perturbation and CRISPR library Cores associated with this U19 application. Subsequent functional analysis of candidate regulatory proteins will be performed in collaboration with other investigators on this grant. The cumulative result of these efforts will be a series of novel gene sets that should define pathways and processes that explain numerous aspects of DC biology as they relate to host defense.

哺乳动物先天免疫系统的树突状细胞（DC）负责检测微生物 遭遇和炎症和适应性免疫的启动。 DC在宿主生理学中的中心作用 吸引了很多关注，这些细胞是主动研究的主题。尽管赞赏 DC的重要性，我们对它们的机制的了解仍然存在很大且根本的差距 规定。例如，DC编码几个检测细菌脂多糖（LPS）的受体，但 仅对一个（TLR4）的功能进行了广泛的检查。我们和其他人的最新工作已经确定 LPS受体CD14和CASPASE-11作为能够诱导继续进行的新信号通路 TLR4（CD14）上游或与TLR4平行（caspase-11）平行。 CD14，TLR4和 caspase-11对于直流激活和宿主防御很重要，但它们的作用机制却很差 定义。因此，我们的知识存在很大的差距，以解释与DC相互作用的最早阶段 细菌。我们还缺乏对DC在感染后期与T细胞相互作用如何与T细胞相互作用的理解，但它是 清楚地表明，这些相互作用会导致两种细胞类型的活性发生深刻的变化。大部分研究 这些相互作用的重点是DC如何促进T细胞活动的变化。相比之下，我们有一个 对T细胞如何影响DC的活性的最低了解。在此应用程序中，我们建议填写这些 通过对小鼠的新调节因素的正向遗传筛查，我们对DC生物学知识的差距。 基于CRISPR的技术提供的基因组编辑的最新进展促进了这种方法。一个 基因发现的管道将通过使用已建立和新兴FACS的渠道生成 测定。这些测定将在体内和Ex Vivo中用于识别DC不足的DC 与微生物或T细胞的相互作用。所有屏幕都将取决于 与此U19应用相关的信息，鼠标扰动和CRISPR库核心。随后的 候选调节蛋白的功能分析将与其他研究者合作进行 在这笔赠款上。这些努力的累积结果将是一系列新型基因集，应定义 解释与宿主防御有关的DC生物学许多方面的途径和过程。