A Conserved Leucine Zipper Motif in Gammaherpesvirus ORF52 Is Critical for Distinct Microtubule Rearrangements.
A Conserved Leucine Zipper Motif in Gammaherpesvirus ORF52 Is Critical for Distinct Microtubule Rearrangements.
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DOI：
10.1128/jvi.00304-17
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发表时间：
2017-09-01
影响因子：
5.4
通讯作者：
Kedes DH
中科院分区：
文献类型：
--
作者：
Loftus MS;Verville N;Kedes DH
生产性病毒感染往往取决于操纵的细胞骨架。疱疹病毒，包括恒河猴rhadinovirus（RRV）及其同源物，致癌的人γ疱疹病毒卡波西肉瘤相关疱疹病毒/人疱疹病毒8（KSHV/HHV 8），利用基于微管（MT）的逆行运输将其基因组递送到细胞核。随后，在生命周期的裂解阶段，成熟的病毒颗粒经历精心安排的易位到细胞质内的专门区域，导致被膜化，二次扩散，然后排出。因此，我们推测RRV可能会在感染的早期和晚期引起细胞骨架的变化。使用共聚焦成像，我们发现，RRV感染导致增厚和乙酰化的MT从MT组织中心（MTOC）后不久，病毒进入和更明显的和弥漫性的MT重组裂解基因表达和病毒体生产的高峰期。我们随后确定了开放阅读框52（ORF 52），一个多功能和丰富的被膜蛋白，作为唯一的病毒编码的组件负责这些细胞骨架的变化。突变和建模分析表明，一个进化上保守的，截短的亮氨酸拉链基序附近的N端，以及一个严格保守的精氨酸残基对ORF 52的C端发挥关键作用，在其重新排列的MT细胞骨架的架构的能力。总之，我们的研究结果与先前描述ORF 52不同作用的研究数据相结合，表明它可能与不同的细胞成分结合，从而允许功能的上下文依赖性调节。重要性对病毒感染过程的透彻理解包括病毒如何操纵其细胞内环境（包括细胞骨架）的知识。例如，改变肌动蛋白或MT聚合的动力学是病毒采用的一种常见策略，以确保有效的进入、成熟和排出，以及通过隔离关键细胞因子来避免抗病毒防御。我们发现，感染RRV，同源的人类病原体KSHV，导致核周包裹乙酰化MT束，并确定ORF 52作为病毒蛋白的基础这些变化。值得注意的是，进入的病毒体能够提供足够的ORF 52以诱导MT增厚和MTOC附近的乙酰化，潜在地帮助将病毒基因组递送到细胞核。虽然在感染的晚期阶段MT改变的功能需要进一步研究，但ORF 52与α疱疹病毒VP 22具有功能和结构相似性，强调了MT细胞骨架操纵对该病毒家族的进化重要性。
Productive viral infection often depends on the manipulation of the cytoskeleton. Herpesviruses, including rhesus monkey rhadinovirus (RRV) and its close homolog, the oncogenic human gammaherpesvirus Kaposi's sarcoma-associated herpesvirus/human herpesvirus 8 (KSHV/HHV8), exploit microtubule (MT)-based retrograde transport to deliver their genomes to the nucleus. Subsequently, during the lytic phase of the life cycle, the maturing viral particles undergo orchestrated translocation to specialized regions within the cytoplasm, leading to tegumentation, secondary envelopment, and then egress. As a result, we hypothesized that RRV might induce changes in the cytoskeleton at both early and late stages of infection. Using confocal imaging, we found that RRV infection led to the thickening and acetylation of MTs emanating from the MT-organizing center (MTOC) shortly after viral entry and more pronounced and diffuse MT reorganization during peak stages of lytic gene expression and virion production. We subsequently identified open reading frame 52 (ORF52), a multifunctional and abundant tegument protein, as being the only virally encoded component responsible for these cytoskeletal changes. Mutational and modeling analyses indicated that an evolutionarily conserved, truncated leucine zipper motif near the N terminus as well as a strictly conserved arginine residue toward the C terminus of ORF52 play critical roles in its ability to rearrange the architecture of the MT cytoskeleton. Taken together, our findings combined with data from previous studies describing diverse roles for ORF52 suggest that it likely binds to different cellular components, thereby allowing context-dependent modulation of function. IMPORTANCE A thorough understanding of the processes governing viral infection includes knowledge of how viruses manipulate their intracellular milieu, including the cytoskeleton. Altering the dynamics of actin or MT polymerization, for example, is a common strategy employed by viruses to ensure efficient entry, maturation, and egress as well as the avoidance of antiviral defenses through the sequestration of key cellular factors. We found that infection with RRV, a homolog of the human pathogen KSHV, led to perinuclear wrapping by acetylated MT bundles and identified ORF52 as the viral protein underlying these changes. Remarkably, incoming virions were able to supply sufficient ORF52 to induce MT thickening and acetylation near the MTOC, potentially aiding in the delivery viral genomes to the nucleus. Although the function of MT alterations during late stages of infection requires further study, ORF52 shares functional and structural similarities with alphaherpesvirus VP22, underscoring the evolutionary importance of MT cytoskeletal manipulations for this virus family.