Varicella Zoster Virus Latency in Human Ganglia

水痘带状疱疹病毒(VZV)是一种普遍存在的人类病原体。原发感染导致儿童水痘，导致脑神经、背根和自主神经节的潜伏期。VZV的重新激活在老年人和免疫功能低下的人群中经常发生，会导致严重的神经系统疾病。带状疱疹和带状疱疹后神经痛占主导地位，但许多人会出现脊髓炎、节段性运动无力、颅神经麻痹和严重的、经常的 脑部出现致命的血管病变。我们还表明，在没有皮疹的情况下，VZV的重新激活可以产生慢性剧烈疼痛。尽管控制VZV潜伏期的机制尚不清楚，但它很可能依赖于病毒基因的转录，从而为我们的假设提供了理论基础，即VZV基因在潜伏期在人类神经节中的表达具有维持潜伏感染的功能。识别潜伏期间表达的VZV基因对于了解潜伏感染至关重要。在高通量阵列技术出现之前，这样的分析是不可行的，并且已经分析了VZV基因组中20%的潜伏病毒转录本。此外，仅仅识别潜伏转录的VZV基因是不够的。由于VZV潜伏期的控制可能涉及病毒蛋白，因此潜伏表达的病毒蛋白的特征对于了解病毒基因调控也是至关重要的。因此，我们的长期目标是对表达的VZV基因进行详细的表征，包括功能分析 在潜伏感染的人类神经节中。我们的具体目标是：(1)通过转录阵列分析鉴定潜伏转录的VZV基因，通过测序确认转录本的真实性，并通过荧光定量(实时)RT-PCR确定病毒转录本的丰度；(2)构建高亲和力表位标记的重组抗体，通过原位免疫组织化学在人神经节切片中共定位潜伏表达的VZV蛋白；以及(3)使用双杂交系统启动蛋白质功能分析，以鉴定和定位蛋白质之间的相互作用。我们将从VZV IE63开始，它是迄今为止在潜伏期检测到的最流行和最丰富的病毒转录本。深入了解VZV基因在潜伏感染的人类神经节中的表达和功能将导致建立潜伏病毒基因调控的可测试模型(在发展中的猴水痘病毒模型中)，并旨在减少与重新激活这种高度嗜神经的人类病原体相关的发病率和死亡率。

Varicella zoster virus (VZV) is an ubiquitous human pathogen. Primary infection causes childhood chickenpox, leading to latency in cranial nerve, dorsal root and autonomic ganglia. VZV reactivation, frequent in the elderly and immunocompromised population, results in significant neurological disease. Zoster and postherpetic neuralgia predominate, but many humans develop myelitis, segmental motor weakness, cranial nerve palsies, and a severe, often fatal vasculopathy in the brain. We have also shown that VZV reactivation can produce chronic intense pain in the absence of rash. Although the mechanism controlling VZV latency is not understood, it is likely to depend upon virus gene transcription, thus providing the rationale for our hypothesis that VZV gene expression in human ganglia during latency functions to maintain latent infection. Identification of the VZV genes expressed during latency is critical to understanding latent infection. Until the advent of high throughput array-based technology, such analyses were not feasible, and <20% of the VZV genome has been analyzed for latent virus transcripts. Further, merely identifying the latently transcribed VZV genes is insufficient. Since control of VZV latency is likely to involve virus proteins, characterization of latently expressed viral proteins is also critical to understanding virus gene regulation. Thus, our long-term goal is the detailed characterization, including functional analysis, of VZV genes expressed in latently infected human ganglia. Our specific aims will: (1) identify latently transcribed VZV genes by transcriptional array analysis, confirm the authenticity of the transcripts by sequencing, and determine the abundance of the virus transcripts by fluorescence-based quantitative (real-time) RT-PCR; (2) construct high affinity epitope-tagged recombinant antibodies to colocalize latently expressed VZV proteins by in situ immunohistochemistry in sections of human ganglia; and (3) initiate protein function analysis using 2-hybrid systems to identify and map protein-protein interactions. We will begin with VZV IE63, the most prevalent and abundant virus transcript detected to date during latency. An in-depth understanding of VZV gene expression and function in latently infected human ganglia will lead to testable models of latent virus gene regulation (in the developing simian varicella virus model) and to therapies designed to reduce morbidity and mortality associated with reactivation of this highly neurotropic human pathogen.