Type III interferon-mediated effector functions of adaptive NK cells involved in control of HCMV infection

自然杀伤（NK）细胞是对抗急性病毒感染的第一道防线的一部分。最近，一种新型的病毒特异性NK细胞被鉴定，其在急性巨细胞病毒（CMV）感染期间扩增并有效介导抗体依赖性细胞毒性（ADCC）。与传统的NK细胞相比，适应性NK细胞可以持续多年，因此有可能有助于长期保护免受再感染或从潜伏期重新激活。这些适应性（也称为记忆）NK细胞的标志是刺激性受体NKG2C的表达，其结合人白细胞抗原E（HLA-E）。除了明确定义的HLA-E/NKG2C轴之外，目前还不清楚哪些其他因素控制病毒感染期间适应性NK细胞的活化。在这方面，我们已经获得了初步数据，显示适应性NK细胞由于IFN 1受体（IFN 1R）的特异性上调而对III型干扰素（即IFN λ（IFN 1）1 - 4）独特地应答。Lambda IFN是最近描述的一类IFN，与经典的I型IFN（IFNalpha/beta）具有同源性，但具有更集中的作用模式，包括针对病毒感染的上皮细胞的一线免疫反应。考虑到适应性NK细胞对作用于常规NK细胞的其他关键细胞因子（例如IL-12和IL-18）的充分描述的低敏感性，适应性NK细胞对IFN 1的响应性是特别相关的。另外的转录组学分析的结果表明，暴露于IFNl在适应性NK细胞中启动高度特异性的病毒防御程序。在本项目中，我们想彻底的特点III型干扰素如何调节适应性NK细胞的效应功能。我们想要理解IFN 1介导的刺激如何通过HLA-E/NKG2C轴与共激活协同作用，并且将使用与抗原加工相关的转运蛋白（TAP）缺陷型HLA-E转染子，其能够通过加载确定的肽来微调HLA-E表面表达。其次，我们将使用转录组学、蛋白质组学和表观遗传学分析来表征IFN 1处理的原代适应性NK细胞中的全局表达程序的变化。第三，我们计划分析IFN 1如何在基于NK细胞与人巨细胞病毒（HCMV）感染的间充质干细胞的共培养的体外感染模型中调节适应性NK细胞的功能。最后，我们将通过分析在化疗和造血干细胞移植（HSCT）后经历HCMV再活化的白血病患者来评估IFN 1介导的适应性NK细胞刺激的临床相关性。总之，拟议的工作预计将导致更好地了解病毒感染期间III型IFN介导的适应性NK细胞的调节，我们希望这将有助于释放其在HCMV临床再激活背景下的细胞治疗中的高翻译潜力。

Natural killer (NK) cells are part of the first line of defense against acute viral infections. Recently, a novel type of virus-specific NK cell was identified that expands during acute cytomegalovirus (CMV) infection and efficiently mediates antibody-dependent cellular cytotoxicity (ADCC). In contrast to conventional NK cells, adaptive NK cells can persist for many years and thus have the potential to contribute to long-term protection against reinfection or reactivation from latency. A hallmark of these adaptive (also known as memory) NK cells is the expression of the stimulatory receptor NKG2C, which binds to human leucocyte antigen E (HLA-E). Besides the well-defined HLA-E/NKG2C axis, it is presently unclear which other factors control the activation of adaptive NK cells during viral infections. In this regard, we have acquired preliminary data showing that adaptive NK cells are uniquely responsive to type III interferons, i.e. IFN lambda (IFNl) 1-4 due to specific up-regulation of the IFNl receptor (IFNlR). Lambda IFNs are a more recently described class of IFNs with homology to the classic type I IFNs (IFNalpha/beta) but with a more focused mode of action, including front-line immune responses against virus-infected epithelial cells. The responsiveness of adaptive NK cells to IFNl is particularly relevant given the well-described low sensitivity of adaptive NK cells to other key cytokines acting on conventional NK cells, such as IL-12 and IL-18. The results of additional transcriptomic analyses suggest that exposure to IFNl initiates a highly specific virus defense program in adaptive NK cells. In the present project, we would like to thoroughly characterize how type III interferons modulate the effector functions of adaptive NK cells. We would like to understand how IFNl-mediated stimulation synergizes with coactivation via the HLA-E/NKG2C axis and will use transporter associated with antigen processing (TAP)-deficient HLA-E transfectants enabling fine-tuning of HLA-E surface expression by loading defined peptides. Second, we will use transcriptomic, proteomic, and epigenetic analyses to characterize the changes in global expression programs in IFNl-treated primary adaptive NK cells. Third, we plan to analyze how IFNl modulates the function of adaptive NK cells in an in vitro infection model based on coculture of NK cells with human cytomegalovirus (HCMV)-infected mesenchymal stem cells. Finally, we will assess the clinical relevance of IFNl-mediated stimulation of adaptive NK cells by analyzing leukemia patients experiencing HCMV reactivation following chemotherapy and hematopoietic stem cell transplantation (HSCT). Together, the proposed work is expected to lead to a better understanding of type III IFN-mediated regulation of adaptive NK cells during virus infections, which we hope will help to unleash their high translational potential in cellular therapy in the setting of clinical reactivation of HCMV.