Molecular study of mouse viral resistance mechanisms

描述（由申请人提供）：本提案基于最近的实验结果，对小鼠CMV感染细胞具有不同的缺失自身MHC-I Dk识别的小鼠配对同源和转基因菌株，以研究NK细胞在病毒感染动物中的作用。该研究项目的长期目标是了解NK细胞中的抑制性受体信号，以及对NK细胞快速响应和杀死病毒感染细胞的能力的影响。该研究提出了一个有趣的问题，即NK抑制受体对病毒感染的识别是否仅仅允许激活NK细胞中的受体信号，还是直接激活NK细胞中有效效应反应和病毒抗性所需的信号级联。这种区别很重要，因为NK细胞对其他免疫细胞有深远的影响，包括启动病毒特异性T细胞效应。研究建议中的一个指导假设是，NK抑制受体对病毒感染的识别传递了一个强有力的信号，允许或“激活”快速NK反应，立即提供保护，防止进一步感染，并开始培养其他免疫细胞，以启动适应性免疫反应。提出了三个具体目标：目标1。将确定抑制受体信号和缺失自身MHC I类识别在NK细胞介导的病毒感染抵抗和增强适应性病毒免疫中的作用。我们将验证ly49g缺失小鼠MCMV感染细胞中缺失自身MHC-I Dk识别需要一种抑制受体的假设。拟议的实验将研究Ly49G2缺乏对先天病毒抗性、病毒特异性T细胞效应物的启动、病毒持久性和死亡率的影响。目标2。将评估抑制受体信号和病毒感染靶细胞缺失自我识别对NK细胞膜近端信号事件的影响，并确定关键的NK细胞表达差异，这可能与先天病毒抗性有因果关系。我们将在病毒感染的背景下研究Ly49G2抑制受体信号，并假设MHC-I Dk感染细胞缺乏自我识别允许或激活NK细胞中以不同生化变化为标志的有效信号，以促进NK细胞的反应性。目标3。将对非mhc基因座进行精细的基因定位、鉴定和表征，这些基因座可以增强NK细胞上的抑制性受体表达和NK细胞介导的病毒抗性。我们将验证Ly49G2受体表达修饰剂和MCMV抗性决定NK细胞对感染的反应、识别感染细胞的能力以及对适应性免疫的影响的假设。NK细胞对人体健康至关重要。它们可以防止恶性肿瘤和病毒感染。NK细胞能识别并应答许多不同类型的病毒。然而，一些病毒能够逃避NK细胞的检测（例如疱疹病毒），而另一些病毒（例如艾滋病毒）能够比NK细胞存活更长时间，智胜或击败NK细胞。因此，了解NK细胞如何识别体内病毒感染的细胞是至关重要的。这一领域的最新进展具有显著的前景，因为我们现在知道NK细胞可以使用直接识别病毒感染细胞的激活受体，从而充分靶向NK细胞杀伤。然而，具有活化受体的NK细胞不能抑制所有的病毒感染。尽管如此，NK细胞也使用抑制性受体来识别和响应病毒感染。不幸的是，一个建立的模型来研究NK抑制受体在病毒感染中的作用之前是不可用的。我们的实验室已经成功地产生了这样一个创新的模型，本研究计划将使用它来研究NK细胞抑制受体的影响及其对抗病毒NK细胞效应功能的贡献。

DESCRIPTION (provided by applicant): This proposal is based on recent experimental results with paired congenic and transgenic strains of mice with divergent missing-self MHC-I Dk recognition of murine CMV infected cells to examine the role of NK cells in virus infected animals. The broad long-term objective for the research project seeks to understand inhibitory receptor signaling in NK cells and the impact on the NK cell's ability to rapidly respond to and kill virus infected cells. An interesting question raised by the research proposal is whether NK inhibitory receptor recognition of virus infection simply permits activation receptor signals in NK cells or directly activates a signal cascade in NK cells needed in efficient effector responses and viral resistance. This distinction is important since NK cells have a profound effect on other immune cells, including the priming of virus-specific T cell effectors. A guiding hypothesis in the research proposal is that NK inhibitory receptor recognition of virus infection delivers a potent signal which permits or 'activates' rapid NK responses to immediately provide protection against further infection and begin to cultivate other immune cells needed to prime adaptive immune responses. Three specific aims are proposed: Aim 1. will determine the effect of inhibitory receptor signaling and missing-self MHC class I recognition of virus infection in NK cell-mediated resistance and enhancement of adaptive viral immunity. We will test the hypothesis that an inhibitory receptor is required in missing-self MHC-I Dk recognition of MCMV infected cells in Ly49G-null mice. Proposed experiments will examine the effect of Ly49G2 deficiency on innate virus resistance, priming of virus-specific T cell effectors, viral persistence and mortality. Aim 2. will assess the effect of inhibitory receptor signaling and missing-self recognition of virus infected target cells on membrane proximal signaling events in NK cells and identify key NK cell expression differences, which may be causally linked with innate virus resistance. We will investigate Ly49G2 inhibitory receptor signaling in the context of virus infection and the hypothesis that missing-self recognition of MHC-I Dk infected cells permits or activates a potent signal, marked by distinct biochemical changes in NK cells, to promote NK cell responsiveness. Aim 3. will perform refined genetic mapping, identification and characterization of non-MHC loci that enhance inhibitory receptor expression on NK cells and NK cell-mediated virus resistance. We will test the hypothesis that modifiers of Ly49G2 receptor expression and MCMV resistance determine how NK cells respond to infection, their capacity to recognize infected cells and consequently their impact on adaptive immunity. Relevance NK cells are vital to human health. They protect against malignancy and virus infection. NK cells recognize and respond to many different types of viruses. However, some viruses are able to evade NK cell detection (e.g. herpesviruses) and others (e.g. HIV) are able to outlast, outwit or defeat NK cells. Thus, it is critical to understand how NK cells recognize virus infected cells in the body. Recent advances in this field have remarkable promise since we know now that NK cells can use activation receptors that directly recognize virus infected cells and therefore target NK cell killing adequately. However, NK cells with activation receptors cannot restrain all virus infections. Despite this, NK cells use inhibitory receptors also to recognize and respond to viral infection. Unfortunately, an established model to investigate the effect of NK inhibitory receptors in viral infections was not available before. Our laboratory has succeeded in generating such an innovative model and this research proposal will use it to investigate the impact of NK cell inhibitory receptors and their contribution to antiviral NK cell effector functions.