I型单纯疱疹病毒（HSV-1）是人群中极为常见的病毒，可引起多种疾病，已成为严重危害人类健康的重要病原之一，目前尚无有效的治疗药物和疫苗。UL2是HSV-1复制过程中一个非常重要的蛋白，单独转染时主要定位于细胞核，但是关于UL2的准确定位、转运机制、定位信号及其生物学功能尚不清楚。本项目拟采用病毒感染及构建UL2荧光蛋白表达质粒来确定UL2的准确定位；构建一系列UL2突变体来确定UL2的核定位信号（NLS）和核输出信号（NES）；通过Ran-GTP、importinα/β突变体和leptomycin B药物抑制实验及免疫共沉淀实验，探讨该蛋白的核质转运机制；运用异源核融合技术研究UL2能否进行核质穿梭；进一步构建含UL2 NLS和NES突变的病毒，研究它们在病毒复制中的作用，从而为阐明UL2在HSV-1感染中的生物学功能奠定基础，也为进一步的抗病毒药物靶点设计和疫苗研制提供理论依据。

Herpes simplex virus 1 (HSV-1), a prevalent human virus that can cause many diseases, has become one of the important pathogen that is severely harmful to human health. However, no effective drug or vaccine has been developed thus far. UL2, a protein that plays vital roles in the process of HSV-1 DNA replication, has been previously shown to target predominantly to the nuclei of transfected cells using indirect immunofluorescence assay (IFA). However, neither the accurate subcellular localization, subcellular localization signals, mechanisms of its subcellular localization and transport, nor its biological function was well understood. . In this study, IFA was firstly conducted to investigate the exact subcellular localization of UL2 in HSV-1-infected cells. To further investigate the subcellular distribution of UL2 in transfected living cells, enhanced yellow fluorescent protein (EYFP)-tagged UL2 variants and fluorescence microscopy were applied. Then, the functional nuclear localization signal (NLS) and nuclear export signal (NES) were mapped and identified. . To explore the nuclear import mechanisms of UL2, a dominant negative (DN) RanGTP (Ran-Q69L), which is deficient in GTP hydrolysis, was introduced to determine whether Ran is required for the nuclear transport of UL2. Further, to identify the cellular receptor responsible for UL2 nuclear targeting and further characterizing the nuclear import pathway of UL2, DN importin α and DN importin β, which lack the ability to bind importin β and Ran respectively, were also introduced to determine whether they are required for the nuclear transport of UL2. To investigate the contribution of CRM1 pathway to the nuclear export of UL2 and to determine whether UL2 is a nucleocytoplasmic shuttling protein, COS-7 cells were transfected with UL2-EYFP expressing plasmid. Then, co-immunoprecipitation assay was employed to verify the interactions of UL2 with cellular receptor importin α, importin β and/or CRM1. Subsequently, heterokaryon assays were performed, with the exception that 3 h prior to and following heterokaryon formation, the cells were treated with leptomycin B to inhibit CRM1 function. . To verify whether the NLS or NES is functional during infection, recombinant viruses with mutations of the NLS and/or the NES in UL2 were constructed, and then IFA was performed to investigate the exact subcellular localization of UL2 in cells infected with wildtype HSV-1 or recombinant virus. To observe whether the subcellular localization of UL2 affects viral replication of HSV-1, the plaque formation and viral proliferation characteristics were researched for each recombinant virus. Together, these jobs are of significance for the further study of the functions of UL2 in HSV-1 infection, as well as for further insights into the design of medicine target of antiviral infection and development of vaccine against HSV-1.