Early events in infection of epithelial cells with Epstein-Barr Virus

EB 病毒感染上皮细胞的早期事件

• 批准号: 7778857
• 负责人: Sarah M Valencia
• 金额: $ 2.66万
• 依托单位: LOUISIANA STATE UNIV HSC SHREVEPORT
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-12-01 至 2011-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7778857
• 关键词: Actins; Address; Antibodies; B-Lymphocytes; Binding; Cell Nucleus; Cells; Complement 3d Receptors; Complement Receptor; Complex; Confocal Microscopy; Cytoplasmic Tail; Development; EBV-associated malignancy; Epithelial; Epithelial Cells; Event; Fluorescent in Situ Hybridization; Glycoproteins; Goals; HIV; Herpesviridae; Homologous Gene; Human; Human Herpesvirus 4; Individual; Infection; Integrins; Intracellular Transport; Laboratories; Lead; Lymphoid; Malignant Neoplasms; Mediating; Microtubules; Oropharyngeal; Phosphorylation; Play; Population; Proteins; Research; Risk; Role; Signal Transduction; Testing; Time; Viral load measurement; Virus; Virus Diseases; Virus Replication; cellular engineering; design; immunosuppressed; inhibitor/antagonist; tumorigenesis

DESCRIPTION (provided by applicant): Epstein-Barr virus (EBV) is an orally transmitted human herpesvirus that is carried by more than ninety percent of the population worldwide and is associated with both lymphoid and epithelial malignancies. The role that it plays in tumorigenesis is complex, but individuals who are immunosuppressed, particularly those infected with the human immunodeficiency virus, are at increased risk of developing EBV-associated malignancies. Part of this increased risk may reflect a loss of control of virus replication and an increase in virus load. The long term goal of the research in the laboratory is to understand how virus spreads and amplifies between and within hosts, moving between the B cells and epithelial cells which are its major targets. Although EBV can bind well to an epithelial cell that lacks CR2 and fusion of an epithelial cell mediated by virus glycoproteins gB and gHgL does not involve CR2, infection of an epithelial cell is much more efficient if CR2 is expressed. Our hypothesis is that this reflects downstream effects on intracellular transport of virus. These are much less well understood than those that occur in a B cell, but can involve interactions between the virus glycoprotein gp350 and the cell protein complement receptor type 2 (CR2 or CD21) and between the virus glycoprotein complex gHgL and the BMRF2 protein and cellular integrins. The immediate goals of the research in this application are to elucidate some of the early events in infection of an epithelial cell and test the hypothesis that virus binding to CR2 triggers important downstream events. These goals will be addressed with three specific aims. Aim one will determine the efficiency with which virus is transported to the nucleus and explore the effects of inhibitors of actin and microtubules. This will be done using a combination of real-time quantitative PCR (RT-QPCR), fluorescence in situ hybridization (FISH) and confocal microscopy. Aim two will determine the role that CR2 plays in increasing efficiency of infection of epithelial cells. This will be done by comparing virus transport to the nucleus in CR2-negative epithelial cells, CR2-positive epithelial cells and cells engineered to express CR2 that lacks a cytoplasmic tail and can no longer interact with FHOS, a formin homolog. Aim three will explore the effects of virus attachment on intracellular signaling. This will be done by analysis of phosphorylation using antibody array analysis. The effects of virus and a soluble form gp350 will be determined. EBV replication is ongoing in the oropharynx of most individuals and may lead to development of epithelial cancers. Understanding how EBV gets into the epithelial cell nucleus and the events that it triggers along the way is important for design of strategies to interrupt or ameliorate the disease that the virus can cause.

描述（由申请人提供）：Epstein-Barr病毒（EBV）是一种口头传播的人疱疹病毒，全球超过90％的人口携带，与淋巴机和上皮恶性肿瘤有关。它在肿瘤发生中所起的作用很复杂，但是免疫抑制的个体，尤其是感染了人类免疫缺陷病毒的人，患有EBV相关性恶性肿瘤的风险增加。这种增加的风险的一部分可能反映了对病毒复制的控制丧失和病毒负荷的增加。实验室研究的长期目标是了解病毒在宿主之间和内部之间如何扩散和放大，在B细胞和上皮细胞之间移动，这是其主要靶标。尽管EBV可以很好地与缺乏CR2的上皮细胞结合，并且由病毒糖蛋白GB和GHGL介导的上皮细胞的融合不涉及CR2，但如果表达CR2，上皮细胞的感染效率更高。我们的假设是，这反映了对病毒细胞内转运的下游影响。这些比在B细胞中发生的知识要多得多，但可能涉及病毒糖蛋白GP350与细胞蛋白质补体2型（CR2或CD21）以及病毒糖蛋白复合蛋白复合GHGL与BMRF2蛋白和细胞整合素之间的相互作用。 该研究中该研究的直接目标是阐明上皮细胞感染的某些早期事件，并检验病毒与CR2结合的假设触发了重要的下游事件。这些目标将以三个特定的目标来解决。 AIM ONE将确定病毒转运到核的效率，并探索肌动蛋白和微管抑制剂的作用。这将使用实时定量PCR（RT-QPCR），荧光原位杂交（FISH）和共聚焦显微镜的组合来完成。目标两个将确定CR2在提高上皮细胞感染效率中所起的作用。这将通过比较CR2阴性上皮细胞，CR2阳性上皮细胞和设计为表达缺乏细胞质尾巴的CR2的细胞中的病毒转运到细胞核的方法来完成，并且无法再与FOMIN同源物（一种福兰）相互作用。 AIM三将探索病毒附着对细胞内信号的影响。这将通过使用抗体阵列分析对磷酸化进行分析来完成。将确定病毒和可溶性形式GP350的作用。大多数个体的口咽中正在进行EBV复制，并可能导致上皮癌的发展。了解EBV如何进入上皮细胞核及其在途中触发的事件对于设计中断或改善病毒可能引起的疾病的策略很重要。