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Regulation of Papillomavirus DNA Replication

乳头瘤病毒 DNA 复制的调控

基本信息

批准号：
6456992
负责人：
MICHAEL R LENTZ
金额：
$ 13.38万
依托单位：
UNIVERSITY OF NORTH FLORIDA
依托单位国家：
美国
项目类别：
财政年份：
2002
资助国家：
美国
起止时间：
2002-05-10 至 2006-04-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/6456992
关键词：
DNA binding protein DNA directed DNA polymerase DNA replication Papillomavirus enzyme activity gene mutation helicase mass spectrometry molecular cloning phosphorylation virus DNA virus protein virus replication

项目摘要

DESCRIPTION (provided by applicant): Cancer of the uterine cervix is one of the few human cancers known to be caused by a specific virus, the human papillomavirus. Despite its known etiology, there are no effective antiviral agents or vaccines to prevent or halt an infection and its progression to cervical cancer. A firm understanding of papillomavirus biology is essential in order to understand specific mechanisms of cancer induction, find methods to prevent infections, and gain insights into the biology of normal and cancer cells. The major objective of the proposed research is to understand mechanisms used by papillomaviruses to regulate their DNA replication. This proposal focuses on the activity of the major viral replication protein, El, and the role of phosphorylation in regulating El function. To fulfill this primary objective, two specific aims will be addressed: one, determine the replication phenotype of phosphorylation defective El mutant viral genomes; and two, analyze purified wild-type and mutant El proteins for biochemical activities and defects. To accomplish these research objectives, a combination of genetic and biochemical approaches will be used. Mutations will be introduced at sites that are phosphorylated on the El protein, and El mutant viral genomes will be analyzed for their replication phenotype in mammalian cells. El proteins that result in replication defects will be expressed in mammalian or insect cells and purified. Wild-type and mutant El proteins will be analyzed for specific El functions, including replication origin DNA binding, helicase enzyme activity, and binding to cellular DNA polymerase. This work will contribute significant new information to the fields of papillomavirus biology and DNA replication, and offer new insights into the unique properties of viral latency.

描述（由申请人提供）：子宫颈癌是一种已知由特定病毒引起的少数人类癌症，人类乳头瘤病毒尽管其病因已知，但没有有效的抗病毒药物预防或阻止感染及其进展的药剂或疫苗，宫颈癌对乳头状瘤病毒生物学的深刻理解对于为了了解癌症诱导的具体机制，找到方法，预防感染，并深入了解正常和癌症的生物学细胞拟议研究的主要目标是了解使用的机制来调节它们的DNA复制该提案的重点是主要病毒复制蛋白El的活性，以及磷酸化调节E1功能。为了实现这一主要目标，将解决两个具体目标：一，确定复制表型磷酸化缺陷El突变病毒基因组;和二，分析纯化的野生型和突变型E1蛋白的生化活性和缺陷。为了实现这些研究目标，遗传和将使用生物化学方法。突变将在在El蛋白上被磷酸化，El突变病毒基因组将被磷酸化。分析它们在哺乳动物细胞中的复制表型。El蛋白质，导致复制缺陷的基因将在哺乳动物或昆虫细胞中表达并被净化将分析野生型和突变型El蛋白的特异性El 功能，包括复制起点DNA结合，解旋酶活性，并与细胞DNA聚合酶结合。这项工作将为以下领域提供重要的新信息：乳头瘤病毒生物学和DNA复制，并提供新的见解，病毒潜伏期的独特特性