Varicella-zoster Virus: Tegument Proteins in Pathogenesis

水痘带状疱疹病毒（VZV）在原发感染期间引起水痘，持续存在于感觉神经节中，并可能从潜伏期重新激活引起带状疱疹。VZV的发病机制取决于其对T细胞、皮肤和感觉神经节的嗜性。VZV疫苗预防水痘和减少带状疱疹的发病率在老年人中是非常有效的。然而，VZV控制可以用第二代重组疫苗改善，该疫苗在皮肤中具有减弱的复制，但对T细胞和神经节的感染性有限。感觉神经节的VZV感染通过从潜伏期再激活而确保其在人群中的存活，从而导致带状疱疹。我们已经建立了一个模型，研究VZV的神经发病机制，使异种移植的人背根神经节（DRG）在小鼠严重联合免疫缺陷（SCID）。我们的VZV皮层/调节蛋白的分析将扩展到检查它们在VZV嗜神经性的功能。我们将专注于IE63，一个重要的立即早期调控蛋白编码的ORF 63，和ORF 66激酶蛋白使用VZV重组有针对性的突变，在这些基因。SCIDhu DRG模型还提供了独特的机会来研究调节病毒基因启动子的细胞反式激活因子如何控制VZV嗜神经性，并确定持续感染的神经元的哪些扰动可能触发VZV再激活。IE63和ORF 66在VZV感染神经节的早期和晚期阶段的作用将在SCIDhu DRG中进行检查。实验将解决四个一般性假设：1）初始VZV复制是神经元中持续VZV DNA拷贝所需的，或影响神经元中持续VZV DNA拷贝的水平; 2）VZ病毒体必须有效地制备和释放; 3）抑制神经细胞凋亡的VZV蛋白促进持久性; 4）DRG的VZV感染反映了与由干扰素（IFN）介导的先天细胞应答的平衡，其优化持久性。VZV基因启动子，像所有疱疹病毒的启动子一样，具有被普遍存在的宿主细胞调节蛋白识别的元件。我们的假设是，细胞蛋白质调节转录的关键ORF 63启动子在初始感染，并需要在感觉神经元的潜伏期过渡。实验，以了解什么细胞应激源可能会诱导VZV从持续感染的DRG再激活将检查热，化学试剂，组蛋白脱乙酰酶抑制剂，干扰神经生长因子信号传导和其他已知的触发器，以提高单纯疱疹病毒再激活。该工作提出了YR。1，Y。2（见特定目的），预计将产生新的信息VZV神经发病机制的分子机制在分化的外周神经元和卫星细胞在人类感觉神经节，并确定选择设计第二代VZV疫苗的遗传变化，已被证明降低毒力在SCIDhu DRG模型在体内。

Varicella zoster virus (VZV) causes varicella during primary infection, persists in sensory ganglia and may reactivate from latency to cause zoster. VZV pathogenesis depends upon its tropisms for T cells, skin and sensory ganglia. VZV vaccines to prevent varicella and to reduce zoster morbidity in the elderly are very effective. However, VZV control could be improved with a 2nd generation recombinant vaccine that has attenuated replication in skin, but limited infectivity for T cells and ganglia. VZV infection of sensory ganglia ensures its survival in the human population through reactivations from latency that result in zoster. We have developed a model for studying VZV neuropathogenesis by making xenografts of human dorsal root ganglia (DRG) in mice with severe combined immunodeficiency (SCID). Our analyses of VZV tegument/regulatory proteins will be extended to examine their functions in VZV neurotropism. We will focus on IE63, an important immediate early regulatory protein encoded by ORF63, and the ORF66 kinase protein using VZV recombinants that have targeted mutations in these genes. The SCIDhu DRG model also offers unique opportunities to investigate how cell transactivators that modulate viral gene promoters may control VZV neurotropism and to identify what perturbations of persistently infected neurons may trigger of VZV reactivation. The roles of IE63 and ORF66 at early and late stages of VZV infection of ganglia will be examined in SCIDhu DRG. Experiments will address four general hypotheses: 1) initial VZV replication is required for, or influences the level of persistent VZV DNA copies in neurons; 2) VZ virions must be made and released efficiently; 3) VZV proteins that inhibit neural cell apoptosis facilitate persistence; 4) VZV infection of DRG reflects an equilibrium with innate cellular responses, mediated by interferons (IFN), which optimizes persistence. VZV gene promoters, like those of all herpesviruses, have elements that are recognized by ubiquitous host cell regulatory proteins. Our hypothesis is that cellular proteins regulate transcription from the critical ORF63 promoter during initial infection and are needed for the transition to latency in sensory neurons. Experiments to understand what cellular stressors may induce VZV reactivation from persistently infected DRG will examine heat, chemical agents, histone deacetylase inhibitors, interference with nerve growth factor signaling and other triggers known to enhance herpes simplex virus reactivation. The work proposed for Yr. 1 and Yr. 2 (see Specific Aims) is expected to yield new information about the molecular mechanisms of VZV neuropathogenesis in differentiated peripheral neurons and satellite cells within human sensory ganglia and to identify options for designing a 2nd generation VZV vaccine with genetic changes that have been proved to reduce virulence in the SCIDhu DRG model in vivo.