Influence of RCMV-encoded vXCL1 on XCR1+ DC

巨细胞病毒(CMV)使用多种策略来调节免疫反应。这些策略之一是表达病毒编码的趋化因子，干扰宿主趋化因子网络，而宿主趋化因子网络在启动和组织白细胞运输方面发挥着重要作用。大鼠CMV的英国和柏林分离株(MuHV-8)编码迄今为止唯一已知的病毒伽马趋化因子。它与内源性大鼠、小鼠和人类γ-趋化因子XCL1的序列相似，因此被命名为病毒XCL1(VXCL1)。为了更好地了解vXCL1的功能并研究与宿主趋化因子的可能竞争，对宿主XCL1的表达以及两种趋化因子与XCR1的结合进行了研究。大鼠XCL1主要由CD8+T细胞、NKT细胞和NK细胞分泌，与人和小鼠的XCL1一样。宿主大鼠XCL1和vXCL1都具有趋化因子样特性，与相应的受体XCR1结合，并仅吸引表达XCR1的CD4-大鼠DC。因此，它们的功能相似。XCR1受体在约80%的脾CD4-XCR1+大鼠DC上表达。病毒XCL1和人类XCL1都是选择性激动剂，分别只激活它们的物种特异性受体rXCR1和hXCR1。相比之下，宿主大鼠XCL1是一种激动剂，可以激活这两种受体。病毒XCL1似乎是一种超级激动剂，与rXCR1的结合能力强于内源性rXCL1趋化因子。为了进一步研究VXCL1和XCR1在啮齿类动物中的相互作用，我们建立了一只XCR1基因敲除大鼠，用于研究病毒感染背景下的受体功能。在目前的提案中，我们将首先描述基因敲除大鼠的特征，并检查其对感染的反应。我们计划研究vXCL1是否只激活大鼠的XCR1，或者是否与不同物种的XCR1存在交叉反应或拮抗活性。进一步，我们打算感染XCR1+DC并分析其感染能力。然后，我们计划用野生型、vxcl1突变型和回复型病毒感染动物，并分析宿主XCL1的表达。我们推测vXCL1通过吸引XCR1+CD4-DC来促进病毒的传播或损害交叉提呈，从而避开CD8+T淋巴细胞所产生的细胞毒性免疫反应。为了检验这一点，我们将确定IE1免疫优势表位的特征，鉴定呈现病毒多肽的表达的MHC等位基因，并最终测试vXCL1是否参与阻止交叉呈递。

Cytomegaloviruses (CMV) use multiple strategies to modulate the immune response. One of these strategies is the expression of virus-encoded chemokines that interfere with the hosts chemokine network which plays an important role in initiating and organizing leukocyte trafficking. The English and Berlin isolates of rat CMV (MuHV-8) encode the so far only known viral gamma-chemokine. Its similar sequence to the endogenous rat, mouse, and human gamma-chemokine XCL1 resulted in its designation as viral XCL1 (vXCL1). To better understand vXCL1 function and investigate a putative competition with the host chemokine, expression of host XCL1 as well as binding of both chemokines to XCR1 was characterized. Rat XCL1 is mostly secreted by CD8+ T cells, NKT cells and NK cells as has been shown for human and mouse XCL1. Both host rat XCL1 and vXCL1 have chemokine-like properties, bind to the corresponding receptor XCR1 and attract exclusively CD4- rat DC that express XCR1. Hence, they function similarly. The receptor XCR1 is expressed on about 80% of splenic CD4- XCR1+ rat DC. Both viral XCL1 and human XCL1 are selective agonists that only activate their species-specific receptors rXCR1 and hXCR1, respectively. In contrast, host rat XCL1 is an agonist that activates both receptors. Viral XCL1 appears to be a super agonist that exhibits superior binding to rXCR1 than the endogenous rXCL1 chemokine. To further study the interaction between vXCL1 and XCR1 in rodents an xcr1 knockout rat was generated to study receptor function in the context of viral infection. In the current proposal, we will initially characterize the knockout rat and examine its response to infection. We plan to investigate if vXCL1 only activates rat XCR1 or if there is cross reactivity or antagonistic activity to XCR1 of different species. Further, we intend to infect XCR1+ DC and analyze their infection capacity. We then plan to infect animals and analyze host XCL1 expression upon infection using wild-type, vxcl1 mutant and revertant viruses. We speculate that vXCL1 attracts XCR1+ CD4- DC to benefit viral dissemination or to impair cross-presentation to circumvent cytotoxic immune responses exerted by CD8+ T lymphocytes. To examine this, we will characterize an IE1 immunodominant epitope, identify an expressed MHC allele presenting a viral peptide and ultimately test if vXCL1 is involved in blocking cross-presentation.