The role of repressive nuclear bodies in latent herpes simplex virus infection

项目摘要/摘要 单纯疱疹病毒1型(HSV-1)是一种普遍存在的人类病原体，在高达90%的 世界人口。HSV-1以潜伏感染的形式在外周神经节持续存在 支配头部和颈部。从潜伏感染中重新激活HSV-1与显著相关 发病率，包括可表现为结膜炎、睑缘炎或角膜上皮炎的疱疹眼病 和间质性角膜炎。反复的眼部病毒再激活可导致复发的疱疹病毒性角膜炎， 仍然是发达国家传染性失明的主要原因。因此，有必要 了解病毒如何在神经元中保持潜伏，最终防止重新激活和复发感染。 在潜伏期间，病毒基因组与抑制性核结构相关，并组装成 异染色质。细胞异染色质相关蛋白在维持HSV-1潜伏期中的作用不是 为人所知。我已经确定了一种细胞蛋白ATRX，它是维持HSV-1潜伏期所必需的。在目标1中 在这项提议中，我将利用一种新的HSV-1潜伏期的初级神经元模型，该模型允许建立 培养在微流室中的外周神经元的潜伏感染。我将测试ATRX是不是 参与维持HSV-1潜伏期的关键细胞蛋白以及ATRX是如何维持的 基于异染色质的基因沉默。此外，我将利用眼部感染模型来理解 体内ATRX耗竭的影响。这些研究将提供对以下贡献的机械性见解 基于异染色质的沉默在维持HSV-1潜伏期和防止反复激活中的作用。 不同病毒基因组的共定位在神经元水平上具有相当大的异质性 核域，可能导致不同类型的潜伏期，或多或少容易重新激活。我 已发现神经元暴露于I型干扰素(干扰素)会导致抑制性PML的形成。 与在没有I型干扰素的情况下建立的潜伏期相比，这种潜伏期的形式更为有限。 因此，在这项提案的目标2中，我将研究PML-核体是如何对I型干扰素做出反应的 并检测PML-核体在压缩潜伏的HSV-1基因组并使其 难以重新激活的。这些目标的最终目的是了解如何长期保存HSV-1 通过维持ATRX与病毒基因组的关联和/或促进HSV- 1抑制性PML核体中的基因组。

PROJECT SUMMARY/ABSTRACT Herpes simplex virus type 1 (HSV-1) is a ubiquitous human pathogen that is present in up to 90% of the world’s population. HSV-1 persists for life in the form of a latent infection in the peripheral ganglia that innervate the head and neck. Reactivation of HSV-1 from latent infection is associated with significant morbidity, including herpetic eye disease which can manifest as conjunctivitis, blepharitis or corneal epithelial and stromal keratitis. Recurrent ocular reactivation of the virus can result in recurrent herpetic keratitis, which remains the leading cause of infectious blindness in the developed world. Therefore, there is a need to understand how the virus remains latent in neurons to ultimately prevent reactivation and recurrent infection. During latency, the viral genome is associated with repressive nuclear structures and is assembled into heterochromatin. The role of cellular heterochromatin-associated proteins in maintaining HSV-1 latency is not known. I have identified one cellular protein, ATRX, that is essential for maintaining HSV-1 latency. In Aim 1 of this proposal, I will utilize a novel primary neuronal model of HSV-1 latency that permits the establishment of a latent infection in peripheral neurons cultured in microfluidic chambers. I will test the hypothesis that ATRX is the key cellular protein involved in preserving HSV-1 latency and examine how ATRX maintains heterochromatin-based gene silencing. In addition, I will utilize an ocular model of infection to understand the effects of ATRX depletion in vivo. These studies will provide mechanistic insights into the contribution of heterochromatin-based silencing in maintaining HSV-1 latency and preventing recurrent reactivation. There is considerable heterogeneity at the neuronal level in the co-localization of viral genomes with different nuclear domains, likely resulting in different types of latency that are more or less susceptible to reactivation. I have found that exposure of neurons to type I interferon (IFN) results in the formation of repressive PML- nuclear bodies, as well as a more restricted form of latency than that established in the absence of type I IFN. Therefore, in Aim 2 of this proposal, I will investigate how PML-nuclear bodies form in response to type I IFN and examine the contribution of PML-nuclear bodies in compacting latent HSV-1 genomes and making them refractory to reactivation. The ultimate goal of these aims is to understand how to preserve long-term HSV-1 latency through either maintaining ATRX association with viral genomes and/or promoting compaction of HSV- 1 genomes in repressive PML-nuclear bodies.