Nucleocapsid envelopment Herpes simplex virus-1

描述（由申请方提供）：这些研究的长期目标是了解疱疹病毒核衣壳蛋白表达的分子机制。所有疱疹病毒最初从受感染细胞的内核膜出芽，这表明从这些研究中获得的信息将适用于这个医学上重要的病毒家族的所有成员。该申请集中于单纯疱疹病毒1 UL 31和UL 34在治疗过程中的作用。基于广泛的初步数据的假设进行了测试。第一种假设提出由UL 31（pUL 3 t）编码的蛋白质与UL 34基因产物（pUL 34）相互作用以介导两种蛋白质正确靶向到内核膜。为了评估体内相互作用的重要性，将测试携带缺乏pUL 34相互作用结构域的UL 31基因的病毒突变体和缺乏pUL 31相互作用区域的UL 34突变体在感染细胞中正确靶向两种蛋白质和介导核衣壳降解的能力。待检验的第二个假设试图理解pUL 31的核纤层缔合的作用。核纤层是排列在核边缘内表面的核细胞骨架或核基质的一部分。一个可能的作用是pUL 31的核纤层缔合介导pUL 31/pUL 34复合物在核边缘的锚定。因此，在瞬时表达测定中，将测试缺乏纤层结合结构域和含有已知的替换这些结构域的纤层结合结构域的UL 31突变体介导pUL 31和pUL 34正确靶向核边缘的能力。然后通过对感染这些病毒的细胞进行电子显微镜检查，评估表达这些蛋白的突变病毒产生包膜病毒体的能力。第二个可能的作用是pUL 31引起部分纤层解聚，以允许核衣壳进入内核膜。为了评估这种可能性，用野生型病毒和缺乏UL 31的缺失病毒感染的细胞层将通过免疫电子显微镜表征。如果野生型病毒感染的细胞含有多孔纤层，而用UL 31突变体感染的细胞不含有多孔纤层，则将探索pUL 31可介导局部纤层解聚的机制。

DESCRIPTION (provided by applicant): The long-term goal of these studies is to understand the molecular mechanisms of herpesvirus nucleocapsid envelopment. All herpesviruses bud initially from the inner nuclear membranes of infected cells suggesting that information gained from these studies will be applicable to all members of this medically important virus family. The application focuses on the role of herpes simplex virus 1 UL31 and UL34 in the envelopment process. Hypotheses based on extensive preliminary data are tested. The first hypothesis proposes that the protein encoded by UL31 (pUL3 t) interacts with the UL34 gene product (pUL34) to mediate proper targeting of both proteins to the inner nuclear membrane. To assess the importance of the interaction in vivo, viral mutants bearing UL31 genes that lack pUL34 interacting domains, and UL34 mutants that lack pUL31 interacting regions will be tested for their abilities to correctly target both proteins in infected cells and to mediate nucleocapsid envelopment. The second hypothesis to be tested seeks to understand the role of nuclear lamina association of pUL31. The nuclear lamina is part of the nucleocytoskeleton or nuclear matrix that lines the inside surface of the nuclear rim. One possible role is that nuclear lamina association of pUL31 mediates anchoring of the pUL31/pUL34 complex at the nuclear rim. Therefore UL31 mutants lacking lamina association domains and containing known lamina binding domains replacing these domains will be tested for their abilities to mediate correct targeting of pUL31 and pUL34 to the nuclear rim in transient expression assays. Mutant viruses expressing these proteins will then be assessed for their ability to produce enveloped virions by electron microscopic examination of cells infected with these viruses. A second possible role is that pUL31 causes partial lamina depolymerization to allow nucleocapsids access to the inner nuclear membrane. To assess this possibility, the lamina of cells infected with wild type virus and a deletion virus lacking UL31 will be characterized by immunoelectron microsopy. If the wild type virus infected cells contain a porous lamina whereas cells infected with the UL31 mutant do not, mechanisms by which pUL31 could mediate local lamina depolymerization will be explored.