宿主蛋白Sec61通过LMP1调控EB病毒复制的研究

LMP1作为EB病毒的关键癌蛋白定位于细胞膜和质膜系统，具有信号转导和基因调控作用， 对EBV相关肿瘤发生至关重要。我们从LMP1与宿主相互作用蛋白的筛选入手，通过Co-IP-MS和Co-focal技术筛选并初步确认了与LMP1有相互作用的宿主分子Sec61，证明易感细胞接种EBV 后上调了Sec61的表达。Sec61是内质网中参与蛋白翻译及转运的重要成分，参与多种病毒的生命周期。本项目拟利用Co-IP、丙氨酸扫描、激光共聚焦、免疫印迹和3D活细胞成像示踪等技术，进一步确证LMP1和Sec61的相互作用，鉴定其互作关键区域和氨基酸位点；明晰Sec61调控LMP1胞内外转运影响EBV基因组稳定和病毒蛋白表达的通路；揭示Sec61通过调控LMP1的定位配置和功能发挥进而促进EBV复制的分子机制。本项目的完成有望对EBV潜伏感染机制提供新认识，并为EBV潜伏感染和相关疾病的治疗提供潜在新靶点。

The viral latent membrane protein 1 (LMP1)，acting as a critical oncoprotein of EBV，is located in the cell membrane and plasma membrane system, plays the role of signal transduction and gene regulation during its infection and is essential for the occurrence of EBV-related human cancers. We started with screening of LMP1 interacting with host proteins, identified and confirmed that Sec61 is a novel LMP1-interacting host protein by immunoprecipitation combined with mass spectrum and with confocal laser scanning microscopy. Further, we have proved that the expression of Sec61 was up-regulated when EBV infected susceptible cells. Since Sec61 is an important component involved in protein translation and transportation in the endoplasmic reticulum, and participates in the life cycle of many viruses. This proposed project would to further verify the interaction of LMP1 and Sec61, and explore the key areas and nucleotide sites of the interaction region, to investigate the pathway that Sec61 regulates the intra- and extracellular transport of LMP1 and then affects the stability of EBV genome and viral protein expression, to elucidate the molecular mechanism that Sec61 promotes the replication of EBV by regulating the positioning and functional exertions of LMP1, through employing Co-IP, alanine scanning mutagenesis, confocal laser scanning microscopy, immunoblotting，3D live cell imaging tracing and other techniques. The completion of the project will expected to provide new understanding of the mechanism of EBV latent infection, and provide new potential targets for the treatment of EBV latent infection and related diseases.