Transcriptional mechanisms of natural killer cell responses during mouse cytomegalovirus infection

小鼠巨细胞病毒感染期间自然杀伤细胞反应的转录机制

• 批准号: 10382356
• 负责人: Timothy E O'Sullivan
• 金额: $ 45.35万
• 依托单位: UNIVERSITY OF CALIFORNIA LOS ANGELES
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-05-06 至 2024-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10382356
• 关键词: ATG3 gene; Acquired Immunodeficiency Syndrome; Activated Natural Killer Cell; Adaptive Immune System; Address; Antigens; Apoptosis; Autophagocytosis; Biological Assay; Biological Process; Cell Count; Cell Cycle; Cell Death; Cell Proliferation; Cell Survival; Cells; ChIP-seq; Clinical; Clonal Expansion; Communicable Diseases; Cytomegalovirus; Cytomegalovirus Infections; DNA sequencing; Data; Defect; Dominant-Negative Mutation; Excision; Exhibits; Foundations; Frequencies; Gene Targeting; Generations; Genes; Genetic Transcription; Genomics; Goals; Health; Human; Immune system; Immunization; Individual; Infection; Innate Immune System; Knockout Mice; Lead; Life; Lymphocyte; Lymphocyte Function; Macaca; Malignant Neoplasms; Measures; Memory; Metabolism; Mitochondria; Modeling; Molecular; Murid herpesvirus 1; Mus; Natural Killer Cells; Newborn Infant; Organ Transplantation; Pathway interactions; Patients; Pattern; Phase; Proliferating; Property; Protein Isoforms; Quantitative Reverse Transcriptase PCR; Regulatory Element; Reporter; Reporting; Repression; Risk; Role; Signal Transduction; Specificity; TP53 gene; Testing; Time; Transgenic Mice; Validation; Viral; Virus; Virus Diseases; Work; cancer cell; cdc Genes; epidemiology study; experimental study; immunosuppressed; in vivo; mitochondrial metabolism; next generation; novel; novel therapeutics; novel vaccines; response; transcription factor; transcriptome; transcriptome sequencing; vaccine strategy

Project Summary Natural killer (NK) cells are lymphocytes of the immune system that can detect and kill virally infected cells. Epidemiological studies have shown that immunosuppressed (e.g. cancer, organ transplant, AIDS) patients and newborns display an enhanced risk for health complications associated with human cytomegalovirus (HCMV) infection that can be life-threating. Previous work has shown that mouse cytomegalovirus (MCMV) infection in mice can accurately model HCMV infection, and demonstrated that NK cells are critical for the control of MCMV. Furthermore, NK cells have been shown to have properties of the adaptive immune system such as recall responses, antigen-specificity, and clonal expansion in mice, macaques, and humans. Although our work has made significant progress into elucidating how autophagy and mitophagy lead to the generation of memory NK cells and the survival of proliferating lymphocytes, the upstream regulatory elements of these fundamental biological processes remain poorly understood in lymphocytes in vivo. Our long-term goals seek to identify the relevant transcriptional signals that induce autophagy, mitophagy, and survival of memory NK cells following viral infection. In RNA-sequencing experiments, we have identified Trp73 (p73) as a transcription factor selectively enriched in memory NK cells following murine cytomegalovirus (MCMV) infection. Further experimental validation by qRT-PCR revealed that two dominant isoforms of Trp73 in mice (TAp73 and ΔNp73) display distinct temporal expression patterns in NK cells during MCMV infection. Importantly, the roles of p73 transcriptional isoforms in lymphocyte responses to viral infection in vivo are unknown. Our exciting preliminary findings suggest that ΔNp73 is transiently induced in effector NK cells and is required for clonal expansion and generation of memory NK cells. In contrast, TAp73 expression is sustained in effector and memory NK cells and is dispensable for effector NK proliferation, but critical for the survival and generation of memory NK cells. In Aim 1, we will determine whether TAp73 influences autophagic removal of dysfunctional mitochondria in NK cells to promote the survival of effector NK cells as they transition to memory cells using cutting-edge autophagy and metabolism assays. In Aim 2, we will determine whether ΔNp73 leads to the proliferation of effector NK cells through repression of p53 to shield NK cells from apoptosis or relieve cell cycle repression. In Aim 3, we will perform RNA and ChIP-sequencing experiments to determine the gene targets of p73 in effector NK cells using genomic analyses. In summary, the proposed studies included in this R01 proposal will contribute to our basic understanding of how primary cells induce mitophagy in vivo, while also contributing to novel clinical strategies to enhance the use of adaptive NK cell responses for immunization against infectious disease.

项目摘要 天然杀手（NK）细胞是可以检测和杀死虚拟感染细胞的免疫系统的淋巴细胞。 流行病学研究表明，免疫抑制（例如癌症，器官移植，艾滋病）患者 新生儿显示出与人类巨细胞病毒有关的健康并发症的风险 （HCMV）可能威胁生命的感染。先前的工作表明小鼠巨细胞病毒（MCMV） 小鼠的感染可以准确地对HCMV感染进行建模，并证明NK细胞对 MCMV的控制。此外，已显示NK细胞具有自适应免疫系统的特性 例如，召回反应，抗原特异性和小鼠，猕猴和人类的克隆扩张。虽然 我们的工作已经取得了重大进步，以阐明自噬和线粒体如何导致这一生成 记忆NK细胞和增殖的淋巴细胞的存活，这些淋巴细胞的上游调节元素 在体内淋巴细胞中，基本生物学过程仍然很少了解。我们的长期目标寻求 确定影响自噬，线粒体和记忆生存的相关转录信号NK 病毒感染后的细胞。在RNA测序实验中，我们已将TRP73（p73）确定为A 在鼠巨细胞病毒（MCMV）之后，选择富集在记忆NK细胞中的转录因子（MCMV） 感染。 QRT-PCR进一步的实验验证表明，小鼠TRP73的两个显性同工型 （TAP73和ΔNP73）在MCMV感染期间，在NK细胞中显示出不同的临时表达模式。 重要的是，p73转录同工型在体内对病毒感染的淋巴细胞反应中的作用是 未知。我们令人兴奋 克隆扩张和生成记忆NK细胞所必需的。相反，TAP73表达得以维持 在效应子和记忆NK细胞中，对于效应子NK增殖是可分配的，但对于生存和 记忆NK细胞的产生。在AIM 1中，我们将确定TAP73是否会影响自噬去除 NK细胞中功能障碍的线粒体功能促进效应NK细胞的存活，它们过渡到记忆 使用尖端的自噬和新陈代谢测定的细胞。在AIM 2中，我们将确定ΔNP73是否引导 通过p53表达效应子NK细胞的扩散，以保护NK细胞免受凋亡或救援 细胞周期表达。在AIM 3中，我们将执行RNA和芯片序列实验以确定基因 使用基因组分析的效应NK细胞中p73的靶标。总而言之，拟议的研究包括 R01提案将有助于我们对原代细胞如何在体内诱导线粒体的基本理解，而 还有助于新的临床策略，以增强自适应NK细胞反应的使用进行免疫 反对传染病。