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

基本信息

批准号：
9926225
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
项目类别：
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/9926225
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 cdc Genes epidemiology study experimental study immunosuppressed in vivo mitochondrial metabolism next generation novel novel therapeutics novel vaccines response transcription factor transcriptome transcriptome sequencing

项目摘要

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.

项目概要自然杀伤 (NK) 细胞是免疫系统的淋巴细胞，可以检测并杀死病毒感染的细胞。流行病学研究表明，免疫抑制（例如癌症、器官移植、艾滋病）患者新生儿出现与人类巨细胞病毒相关的健康并发症的风险增加先前的研究表明，小鼠巨细胞病毒（MCMV）感染可能会危及生命。小鼠感染可以准确模拟 HCMV 感染，并证明 NK 细胞对于此外，NK 细胞已被证明具有适应性免疫系统的特性。例如小鼠、猕猴和人类的回忆反应、抗原特异性和克隆扩增。我们的工作在阐明自噬和线粒体自噬如何导致生成方面取得了重大进展记忆 NK 细胞和增殖淋巴细胞的存活，这些细胞的上游调控元件我们对体内淋巴细胞的基本生物学过程仍知之甚少。识别诱导自噬、线粒体自噬和记忆 NK 存活的相关转录信号在 RNA 测序实验中，我们将 Trp73 (p73) 鉴定为病毒感染后的细胞。鼠巨细胞病毒 (MCMV) 后记忆 NK 细胞中选择性富集的转录因子 qRT-PCR 的进一步实验验证表明，小鼠体内有两种主要的 Trp73 亚型。 (TAp73 和 ΔNp73) 在 MCMV 感染期间 NK 细胞中表现出不同的时间表达模式。重要的是，p73转录亚型在体内淋巴细胞对病毒感染的反应中的作用是我们令人兴奋的初步研究结果表明，ΔNp73 在效应 NK 细胞中被短暂诱导，并且是存在的。相比之下，TAp73 的表达是持续的。 NK 细胞存在于效应子和记忆 NK 细胞中，对于效应子 NK 增殖来说是可有可无的，但对于生存和生存至关重要。在目标 1 中，我们将确定 TAp73 是否影响自噬清除。 NK 细胞中功能失调的线粒体可促进效应 NK 细胞在向记忆过渡时的存活在目标 2 中，我们将确定 ΔNp73 是否领先。通过抑制 p53 来抑制效应 NK 细胞的增殖，从而保护 NK 细胞免于凋亡或缓解在目标 3 中，我们将进行 RNA 和 ChIP 测序实验来确定基因。使用基因组分析确定效应 NK 细胞中 p73 的靶点总之，本研究中包含了拟议的研究。 R01提案将有助于我们对原代细胞如何在体内诱导线粒体自噬的基本理解，同时还有助于制定新的临床策略，以增强适应性 NK 细胞反应在免疫中的应用对抗传染病。