Alpha-herpesvirus infection induces the formation of nuclear actin filaments.
Alpha-herpesvirus infection induces the formation of nuclear actin filaments.
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DOI：
10.1371/journal.ppat.0020085
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发表时间：
2006-08
期刊：
影响因子：
6.7
通讯作者：
Enquist LW
中科院分区：
文献类型：
--
作者：
Feierbach B;Piccinotti S;Bisher M;Denk W;Enquist LW
疱疹病毒是在受感染细胞的细胞核中复制的大的双链DNA病毒。通过建立用于浓缩病毒和宿主因子的核隔室来实现宿主核中病毒复制和组装的空间控制。这些分隔区是如何建立和维持的，人们仍然知之甚少。伪狂犬病病毒（PRV）是一种α-疱疹病毒，常用于研究疱疹病毒在神经系统中的入侵和传播。在这里，我们报告，PRV和单纯疱疹病毒1型感染的神经元在细胞核中形成肌动蛋白丝的结果。在未感染细胞的细胞核中没有发现丝状肌动蛋白。核肌动蛋白丝物理上与病毒衣壳相关，如连续块面扫描电子显微镜和共聚焦显微镜所示。使用一个绿色荧光蛋白标记的病毒衣壳蛋白（VP26），我们表明，核肌动蛋白丝形成之前，衣壳组装和所需的有效形成病毒衣壳组装位点。我们发现肌动蛋白聚合动力学（例如，研磨）对于这些位点的形成不是必需的。绿色荧光蛋白-VP26焦点与肌动蛋白马达肌球蛋白V共定位，表明病毒衣壳使用基于肌球蛋白的定向运输沿沿着核肌动蛋白丝行进。肌动蛋白丝的形成需要病毒的转录，而不是病毒DNA的复制。PRV或单纯疱疹病毒1型的感染导致神经元中核肌动蛋白丝的形成，并且PRV感染上皮细胞系导致类似的表型，这一发现证明F-肌动蛋白在疱疹病毒组装中起保守作用。我们的研究结果提出了一种机制，通过该机制，组装结构域被组织在感染的细胞内，并提供深入了解病毒感染周期和宿主肌动蛋白细胞骨架是如何整合的，以促进感染过程。亚细胞组织和运输的调节对于控制关键的生物过程是必不可少的。然而，我们的知识往往受到阻碍，因为这些过程往往是短暂的，难以研究。关于机会微生物如何劫持细胞机制的研究为各种正常细胞过程提供了见解。例如，用细胞内微生物如单核细胞增生李斯特菌、志贺氏菌属、立克次体属，和牛痘病毒，大大增加了我们对肌动蛋白细胞骨架的动态性质的理解。然而，对亚细胞组织和细胞核中货物运输的了解要少得多。作者发现，α-疱疹病毒感染神经元导致细胞核中肌动蛋白丝的瞬时形成。这些细丝并不填充细胞核，而是与新形成的病毒衣壳结合。核肌动蛋白丝最初是在外周神经系统组织中使用新的成像技术，连续切片扫描电子显微镜由Winfried Denk（合著者）开创的。他们的研究结果表明，细胞核肌动蛋白丝的形成是对感染的一般应激反应的一部分，但随后被增选，可能是为了引导衣壳运输到沿着核膜出芽的部位。这项工作阐明了病毒生命周期中一个不太清楚的部分，并为未来研究控制货物如何在细胞核中组织和移动的工作奠定了基础。
Herpesviruses are large double-stranded DNA viruses that replicate in the nuclei of infected cells. Spatial control of viral replication and assembly in the host nucleus is achieved by the establishment of nuclear compartments that serve to concentrate viral and host factors. How these compartments are established and maintained remains poorly understood. Pseudorabies virus (PRV) is an alpha-herpesvirus often used to study herpesvirus invasion and spread in the nervous system. Here, we report that PRV and herpes simplex virus type 1 infection of neurons results in formation of actin filaments in the nucleus. Filamentous actin is not found in the nucleus of uninfected cells. Nuclear actin filaments appear physically associated with the viral capsids, as shown by serial block-face scanning electron micropscopy and confocal microscopy. Using a green fluorescent protein-tagged viral capsid protein (VP26), we show that nuclear actin filaments form prior to capsid assembly and are required for the efficient formation of viral capsid assembly sites. We find that actin polymerization dynamics (e.g., treadmilling) are not necessary for the formation of these sites. Green fluorescent protein-VP26 foci co-localize with the actin motor myosin V, suggesting that viral capsids travel along nuclear actin filaments using myosin-based directed transport. Viral transcription, but not viral DNA replication, is required for actin filament formation. The finding that infection, by either PRV or herpes simplex virus type 1, results in formation of nuclear actin filaments in neurons, and that PRV infection of an epithelial cell line results in a similar phenotype is evidence that F-actin plays a conserved role in herpesvirus assembly. Our results suggest a mechanism by which assembly domains are organized within infected cells and provide insight into how the viral infectious cycle and host actin cytoskeleton are integrated to promote the infection process. Regulation of subcellular organization and transport is essential for control of crucial biological processes. However, our knowledge often is hampered because these processes tend to be transient and difficult to study. Studies of how opportunistic microbes hijack cellular machinery have provided insights into various normal cell processes. For example, studies with intracellular microorganisms, such as Listeria monocytogenes, Shigella spp., Rickettsia spp., and vaccinia virus, have significantly increased our understanding of the dynamic nature of the actin cytoskeleton. However, much less is known about subcellular organization and transport of cargo in the nucleus. The authors have discovered that alpha-herpesvirus infection of neurons leads to the transient formation of actin filaments in the nucleus. These filaments do not fill the nucleus, but rather associate with newly formed viral capsids. The nuclear actin filaments were initially identified in peripheral nervous system tissue using a new imaging technology, serial section scanning electron microscopy pioneered by Winfried Denk (a co-author). Their results suggest that nuclear actin filaments form as part of a general stress response to infection, but then are co-opted, perhaps to direct capsid transport to sites of budding along the nuclear envelope. This work illuminates a less well understood part of the viral life cycle and sets the stage for future work investigating control of how cargo is organized and moved in the nucleus.