Cell Entry and Spread by Polyoma Virus

描述（由申请人提供）：本申请的长期目标是了解小鼠多瘤病毒进入细胞的机制。这种病毒的宿主范围很广。它在其天然宿主的多种细胞类型中复制和诱导肿瘤。具体目标首先是确定某些糖脂是否构成病毒的唯一一类细胞受体，或者某些糖蛋白是否也可以作为功能性受体。这将使用糖脂生物合成缺陷但糖蛋白途径不受影响的小鼠进行。将进行研究，以确定病毒在内质网中解体的步骤和病毒用于进行必要的解体步骤并进入细胞核的细胞因子。在这方面正在研究的因素是氧化还原酶ERp29，蛋白质二硫键异构酶，derlin分子伴侣，和Ran GT3。一个具体的目标是在体外产生和表征被准备用于膜插入和从ER逃逸的改变的颗粒。最后，与哈佛的Xiaowei Zhuang博士的实验室合作，将尝试跟踪活细胞中的单个病毒颗粒，因为它们进入并建立感染。我们将尝试生产多瘤颗粒标记上的微型染色体和外衣壳蛋白与荧光探针，研究这些组件的脱壳和分离。许多病毒进入细胞的过程尚未完全了解。多瘤病毒组包括几种对人类具有致病性的病毒，包括JC、BK和可能的SV40。了解这些病毒进入细胞的途径可能为抗病毒治疗或预防提供机会。

DESCRIPTION (provided by applicant): The long-term objectives of this application are to understand the mechanisms of cell entry by the mouse polyoma virus. This virus has a broad host range. It replicates and induces tumors in a wide variety of cell types in its natural host. The Specific Aims are first to determine whether certain glycolipids constitute the only class of cell receptors for the virus or whether certain glycoproteins may also serve as functional receptors. This will be done using a mouse deficient in glycolipid biosynthesis but unaffected in glycoprotein pathways. Investigations will be carried out to identify the steps in virus disassembly in the endoplasmic reticulum and the cellular factors the virus utilizes to undergo the necessary steps of disassembly and to enter the nucleus. Factors under investigation in this regard are the oxidoreductase ERp29, protein disulfide isomerase, the derlin chaperones, and Ran GTPase. A specific goal is to generate in vitro and characterize the altered particle that is primed for membrane insertion and escape from the ER. Finally, in collaboration with Dr. Xiaowei Zhuang's lab at Harvard, attempts will be made to follow single virus particles in live cells as they enter and establish infection. We will attempt to produce polyoma particles labeled both on the minichromosome and the outer capsid protein with fluorescent probes to study decapsidation and separation of these components. The processes of cell entry by many viruses are not fully understood. The polyoma group includes several viruses that are pathogenic in humans, including JC, BK and possibly SV40. Understanding the pathways of cell entry by these viruses may suggest opportunities for anti-viral therapy or prevention.