Gene Expression in HSV-1 Latency After Corneal Infection

描述（由申请人提供）：1型单纯疱疹病毒（HSV-1）是角膜失明的主要原因。在人类中，角膜疾病是由HSV-1在初次感染时在宿主感觉神经节中建立的潜伏（静止）状态反复重新激活引起的。包括我们在内的几个实验室最近的发现挑战了一般的假设，即在HSV-1潜伏期间缺乏病毒蛋白合成妨碍了对潜伏感染神经元的免疫检测和对潜伏状态的免疫调节。更确切地说，这些发现提出了一种潜伏期的动态模型，在这种模型中，许多重新激活的尝试被潜伏感染的神经节中的CD8+ T细胞的抢先反应所中止。我们的基本假设是，潜伏期间的基因表达允许神经节CD8+T细胞提供“及时”的机制来抑制体内病毒的产生。在本提案中，抗原在潜伏期的表达和增强免疫介导的病毒潜伏期维持的概念将被探讨。特异性目标1将验证基因表达在潜伏期和诱导再激活期间发生在TG中的假设，而不产生感染性病毒和“真正的晚期”病毒基因。我们将从不同的候选病毒启动子中获得调节两种荧光蛋白的重组HSV-1。小鼠将通过角膜途径感染，建立潜伏期。结合共聚焦成像技术，我们将在潜伏感染和再激活诱导小鼠的TG中鉴定HSV-1启动子的活性。我们的模型预测，在缺乏晚期（γ -2）基因和感染性病毒的小鼠潜伏期中，特异性HSV-1启动子是活跃的。在特异性目标2中，我们将验证CD8+ T细胞进入神经节的免疫浸润以及它们免于再激活的保护，可以通过潜伏感染神经元中病毒表位的持续表达来增强。在潜伏活性启动子的作用下，将开发出表达免疫优势肽多聚体的病毒。潜伏感染这些病毒的小鼠将被评估CD8+ T细胞浸润和对诱导再激活的抵抗。这些研究将为疫苗的设计奠定基础，以增强对潜伏期和再激活的免疫调节。

DESCRIPTION (provided by applicant): Herpes simplex virus type 1 (HSV-1) is a major cause of corneal blindness. In humans, corneal disease results from repeated reactivation of HSV-1 from a latent (quiescent) state that was established in the host sensory ganglia at the time of primary infection. Recent findings from several laboratories, including ours, challenge the general supposition that a lack of viral protein synthesis during HSV-1 latency precludes immune detection of latently infected neurons and immune modulation of the latent state. Rather, these findings give rise to a dynamic model of latency in which many reactivation attempts are aborted by a preemptive CD8+ T cell response in the latently infected ganglia. Our underlying hypothesis is that gene expression during latency allows ganglionic CD8+T cells to provide a 'just in time' mechanism to inhibit virus production in vivo. In this proposal, the concepts of antigen expression during latency and the enhancement of the immune-mediated maintenance of viral latency will be explored. Specific Aim 1 will test the hypothesis that gene expression occurs in the TG during latency and induced reactivation without the production of infectious virus and 'true late' viral genes. We will derive recombinant HSV-1 that regulate two fluorescent proteins from different candidate viral promoters. Mice will be infected through the corneal route to establish latency. In conjunction with confocal imaging techniques, we will identify HSV-1 promoter activities in the TG of latently infected and reactivation-induced mice. Our model predicts that specific HSV-1 promoters are active during murine latency in the absence of late (gamma-2) genes and infectious virus. In Specific Aim 2, we will test the hypothesis that immune infiltration of CD8+ T cells into the ganglia, and the protection they afford from reactivation, can be enhanced by persistent expression of viral epitopes in latently infected neurons. Viruses will be developed which express multimers of an immunodominant peptide under latency active promoters. Mice latently infected with such viruses will be assessed for CD8+ T cell infiltration and resistance to induced reactivation. Such studies will establish foundations for the design of vaccines to augment immune regulation of latency and reactivation.