Regulation of NK cell antiviral responses through alternative splicing

通过选择性剪接调节 NK 细胞抗病毒反应

• 批准号: 10258225
• 负责人: Daniel Ramos Ram
• 金额: $ 17.58万
• 依托单位: BETH ISRAEL DEACONESS MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-04-15 至 2024-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10258225
• 关键词: Abnormal Cell; Acute; Affect; Alternative Splicing; Antigens; Antisense Oligonucleotides; Antiviral Agents; Antiviral Response; Area; Cancerous; Candidate Disease Gene; Cell physiology; Cells; Cellular biology; Characteristics; Chronic; Cryopreservation; Cytomegalovirus; Data; Development; Disease; ES04; Effector Cell; Epigenetic Process; Event; Evolution; Exhibits; Exons; Family; Family member; Female; Flow Cytometry; Functional disorder; Gene Expression; Gene Expression Profile; Gene Expression Regulation; Genes; Genetic Transcription; Goals; HIV; HIV Infections; HIV therapy; Human; Immune; Immune response; Immunity; Immunology; Impairment; Inactivated Vaccines; Individual; Infection; Infection Control; Innate Immune Response; Innate Immune System; Laboratories; Lead; Macaca; Macaca mulatta; Malignant Neoplasms; Mediating; Memory; Modification; NUP214 gene; Natural Killer Cells; Output; Pattern; Peptides; Peripheral Blood Mononuclear Cell; Persons; Play; Population; Property; Protein Isoforms; Publishing; RNA; RNA Splicing; Regulation; Reporting; Research; Resistance; Role; SIV; Sampling; Small Interfering RNA; Spliced Genes; TNFRSF6 gene; Testing; Therapeutic Intervention; Third Generation Sequencing; Tissues; Training; Transcript; Vaccination; Vaccines; Variant; Viral; Viral reservoir; Virus; Virus Diseases; Work; acute infection; adaptive immunity; antiretroviral therapy; chronic infection; cohort; environmental change; epitranscriptomics; gene product; immunosenescence; improved; in vivo; influenza virus vaccine; insight; interest; male; member; novel; peripheral blood; post-doctoral training; pre-doctoral; protein complex; receptor; response; stem; transcription factor; transcriptome; transcriptome sequencing; transcriptomics; tumor; vaccine response

Project Summary/Abstract Natural killer (NK) cells have been shown to play a significant role in regulating viral infections as well as in eliminating cancerous cells. Traditionally, NK cells are classified as members of the innate immune system because of their ability to respond rapidly and non-specifically to infected or abnormal cells. Interestingly, burgeoning evidence suggests NK cells may also possess peptide-specific memory responses (previously only attributable to adaptive immunity), especially in the context of several viral infections including HIV/SIV and CMV. How NK cells can form these adaptive responses is the subject of an emerging area in immunology research, which also includes studying trained immunity of traditionally innate effectors like NK cells. Whether adaptive NK cell responses are the result of epigenetic modifications as observed in trained immunity, or the product of pre- formed polymorphic receptors like KIRs or members of the NKG2 family which interact with MHC molecules, or yet to be discovered receptors is to be determined. Here, we propose that investigating the NK cell transcriptome can provide insights into how infection or disease states can modulate NK cell function particularly by assessing how changes in alternative gene splicing following infection result in NK cell dysfunction or the formation of adaptive NK cell responses. Alternative splicing is a mechanism utilized by cells in order to produce multiple gene products from an individual gene, by splicing exons together in various combinations, sometimes leading to gene products with very different functions. Regulation of gene splicing is complicated, but it has been shown that environmental changes as are seen during viral infection or disease can gene splicing patterns. Using preliminary data from sorted NK cells in an acute SIV-infection cohort we have already identified several candidate alternatively spliced genes that exhibit differential isoform expression following SIV infection. Of particular interest are members of the bHLH transcription factor (TF) family due to their association with the formation of memory responses and NK cell development. Further, in a proof-of-concept study we have shown that NKp30 isoform expression is altered following ex vivo stimulation or even following vaccination in rhesus macaque and in human NK cell samples. Here, I propose to identify novel gene candidates and assess the roles of the bHLH TF splice variants and other candidate genes in rhesus macaque and human NK cells. Through this work I aim to determine whether specific gene isoform expression impacts normal NK cell functions or the formation of adaptive NK cell responses in vivo in rhesus macaques during acute and chronic SIV infection. The approaches outlined in this proposal will allow me to combine my postdoctoral training in NK cell biology with my predoctoral training assessing the role of alternative splicing in vivo in order to determine the roles that several gene candidates have on normal or adaptive NK cell responses during retroviral infection. Our ultimate goal is to identify and target specific isoforms to manipulate NK cell function in vivo for the purposes of improving current vaccine responses or eliciting targeted NK cell-specific therapies for HIV.

项目概要/摘要 自然杀伤 (NK) 细胞已被证明在调节病毒感染以及在 消除癌细胞。传统上，NK 细胞被归类为先天免疫系统的成员 因为它们能够对受感染或异常细胞做出快速且非特异性的反应。有趣的是， 越来越多的证据表明 NK 细胞也可能具有肽特异性记忆反应（以前仅 归因于适应性免疫），特别是在包括 HIV/SIV 和 CMV 在内的几种病毒感染的情况下。 NK 细胞如何形成这些适应性反应是免疫学研究新兴领域的主题， 其中还包括研究 NK 细胞等传统先天效应细胞的训练有素的免疫力。是否自适应NK 细胞反应是在训练有素的免疫中观察到的表观遗传修饰的结果，或者是预免疫的产物。 形成多态性受体，如 KIR 或 NKG2 家族成员，与 MHC 分子相互作用，或 尚未发现的受体尚待确定。在这里，我们建议研究 NK 细胞转录组 可以深入了解感染或疾病状态如何调节 NK 细胞功能，特别是通过评估 感染后替代基因剪接的变化如何导致 NK 细胞功能障碍或形成 适应性 NK 细胞反应。选择性剪接是细胞利用的一种机制，以产生多种 来自单个基因的基因产物，通过以各种组合将外显子剪接在一起，有时导致 具有非常不同功能的基因产物。基因剪接的调控很复杂，但已被证明 病毒感染或疾病期间的环境变化可以影响基因剪接模式。 使用急性 SIV 感染队列中分选的 NK 细胞的初步数据，我们已经确定了几种 SIV 感染后表现出差异亚型表达的候选可变剪接基因。的 特别感兴趣的是 bHLH 转录因子 (TF) 家族的成员，因为它们与 记忆反应的形成和 NK 细胞的发育。此外，在概念验证研究中我们已经表明 在恒河猴中，NKp30亚型表达在离体刺激甚至疫苗接种后发生改变 猕猴和人类 NK 细胞样本中。在这里，我建议确定新的候选基因并评估其作用 恒河猴和人类 NK 细胞中的 bHLH TF 剪接变体和其他候选基因。通过这个 我的工作目的是确定特定基因亚型表达是否影响正常 NK 细胞功能或 急性和慢性 SIV 感染期间恒河猴体内适应性 NK 细胞反应的形成。 本提案中概述的方法将使我能够将我在 NK 细胞生物学方面的博士后培训结合起来 我的博士前培训评估了体内选择性剪接的作用，以确定其作用 一些候选基因在逆转录病毒感染期间具有正常或适应性 NK 细胞反应。我们的终极 目标是识别和靶向特定亚型以操纵体内 NK 细胞功能，从而改善 当前的疫苗反应或引发针对 HIV 的靶向 NK 细胞特异性疗法。